Urieli-Shoval S, Gruenbaum Y, Razin A
J Bacteriol. 1983 Jan;153(1):274-80. doi: 10.1128/jb.153.1.274-280.1983.
Two DNA methylase activities of Escherichia coli C, the mec (designates DNA-cytosine-methylase gene, which is also designated dcm) and dam gene products, were physically separated by DEAE-cellulose column chromatography. The sequence and substrate specificity of the two enzymes were studied in vitro. The experiments revealed that both enzymes show their expected sequence specificity under in vitro conditions, methylating symmetrically on both DNA strands. The mec enzyme methylates exclusively the internal cytosine residue of CCATGG sequences, and the dam enzyme methylates adenine residues at GATC sites. Substrate specificity experiments revealed that both enzymes methylate in vitro unmethylated duplex DNA as efficiently as hemimethylated DNA. The results of these experiments suggest that the methylation at a specific site takes place by two independent events. A methyl group in a site on one strand of the DNA does not facilitate the methylation of the same site on the opposite strand. With the dam methylase it was found that the enzyme is incapable of methylating GATC sites located at the ends of DNA molecules.
通过DEAE - 纤维素柱层析对大肠杆菌C的两种DNA甲基化酶活性，即mec（表示DNA - 胞嘧啶甲基化酶基因，也称为dcm）和dam基因产物进行了物理分离。在体外研究了这两种酶的序列和底物特异性。实验表明，两种酶在体外条件下均表现出预期的序列特异性，在两条DNA链上对称甲基化。mec酶仅甲基化CCATGG序列的内部胞嘧啶残基，而dam酶甲基化GATC位点的腺嘌呤残基。底物特异性实验表明，两种酶在体外对未甲基化的双链DNA和半甲基化DNA的甲基化效率相同。这些实验结果表明，特定位点的甲基化是由两个独立事件发生的。DNA一条链上某位点的甲基基团不会促进其互补链上同一位点的甲基化。对于dam甲基化酶，发现该酶无法甲基化位于DNA分子末端的GATC位点。
