Cloning, characterization, and expression in Escherichia coli of the gene coding for the CpG DNA methylase from Spiroplasma sp. strain MQ1(M.SssI).
We describe here the cloning, characterization and expression in E. coli of the gene coding for a DNA methylase from Spiroplasma sp. strain MQ1 (M.SssI). This enzyme methylates completely and exclusively CpG sequences. The Spiroplasma gene was transcribed in E. coli using its own promoter. Translation of the entire message required the use of an opal suppressor, suggesting that UGA triplets code for tryptophan in Spiroplasma. Sequence analysis of the gene revealed several UGA triplets, in a 1158 bp long open reading frame. The deduced amino acid sequence revealed in M.SssI all common domains characteristic of bacterial cytosine DNA methylases. The putative sequence recognition domain of M.SssI showed no obvious similarities with that of the mouse DNA methylase, in spite of their common sequence specificity. The cloned enzyme methylated exclusively CpG sequences both in vivo and in vitro. In contrast to the mammalian enzyme which is primarily a maintenance methylase, M.SssI displayed de novo methylase activity, characteristic of prokaryotic cytosine DNA methylases.
Renbaum
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P
Department of Cellular Biochemistry, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
Abrahamove
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D
Fainsod
A
A
Wilson
G G
GG
Rottem
S
S
Razin
A
A
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D015254:DNA Modification Methylases / Q000235:genetics* / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D015226:Dinucleoside Phosphates / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D013149:Spiroplasma / Q000201:enzymology / Q000235:genetics*; D013379:Substrate Specificity
D015254:DNA Modification Methylases; D004269:DNA, Bacterial; D015226:Dinucleoside Phosphates; C015772:CpG dinucleotide