An orphan DNA (cytosine-5-)-methyltransferase in Vibrio cholerae.
5-Methyl cytosine (m5C) was detected in genomic DNA of the enteric pathogen Vibrio cholerae by HPLC analysis and immunoblotting with m5C-specific antibody. Although cleavage with the restriction endonuclease EcoRII revealed the absence of a Dcm homologue in V. cholerae, analysis of the genome sequence indicated the presence of a gene, designated in this study as vchM, which encodes a DNA (cytosine-5-)-methyltransferase (m5C-MTase) designated M.Vch. M.Vch is not associated with a restriction endonuclease or a mismatch very short patch repair (Vsr)-like endonuclease and is hence an 'orphan' or solitary MTase, although analysis of a phylogenetic tree indicated that related cytosine MTases are all components of restriction-modification systems. M.Vch recognizes and methylates the first 5' C in the degenerate sequence 5'-RCCGGY-3'. RT-PCR analysis suggested that vchM gene expression is increased during the stationary phase of growth. During stationary phase, the spontaneous mutation frequency in the V. cholerae wild-type strain was significantly higher than in the corresponding vchM mutant strain, suggesting that the presence of M.Vch and the absence of a very short patch (VSP) repair-like system imposes upon V. cholerae a mutator phenotype.
Biophysics Division, Indian Institute of Chemical Biology, 4 Raja S. C. Mullick Road, Calcutta 700 032, India.
Sanjib
S
Banerjee
Biophysics Division, Indian Institute of Chemical Biology, 4 Raja S. C. Mullick Road, Calcutta 700 032, India.
Rukhsana
R
Chowdhury
D002851:Chromatography, High Pressure Liquid; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D015151:Immunoblotting; D009154:Mutation; D010802:Phylogeny; D010957:Plasmids / Q000378:metabolism; D012329:RNA, Bacterial / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014158:Transcription, Genetic; D014734:Vibrio cholerae / Q000201:enzymology* / Q000235:genetics
D004269:DNA, Bacterial; D012329:RNA, Bacterial; D012333:RNA, Messenger; D004248:DNA (Cytosine-5-)-Methyltransferases