[Fractionation and purification of DNA methylases and specific endonucleases from cells of Escherichia coli SK].
Fractionation and purification of DNA methylases and specific endonucleases from E. coli SK responsible for DNA specificity to host prokaryotic cells were studied. The most efficient purification was achieved by precipitation of proteins by 0.6 saturated ammonium sulfate with subsequent chromatography on KM-cellulose and concentration of fractions by dialysis against glycerol. Under these conditions the methylase activity produced 4 discrete fractions. Due to purification the specific activity of methylases increased 11--20-fold in various fractions. Methylase from the first (A) and fourth (BII) peaks catalyzed the methylation of cytosine to produce 5-methylcytosine; methylase from the third peak (BI) methylated adenine to produce 6-methylaminopurine. The chemical specificity of the second peak (B) methylase could not be established due to very high lability of the enzyme in this fraction. Specific endonuclease was found in the gradient zones eluted by 0.1--0.2 M and 0.65--0.75 M NaCl. It is assumed that those enzymes providing for DNA hydrolysis up to the formation of high--molecular discrete fragments, are restricting endonucleases of the SK system. The results obtained strongly suggest the existence of several types of methylases and restricting endonucleases in E. coli SK cells.
D004247:DNA / Q000378:metabolism; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000302:isolation & purification* / Q000378:metabolism; D004262:DNA Restriction Enzymes / Q000032:analysis; D004720:Endonucleases / Q000302:isolation & purification* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology*; D006868:Hydrolysis; D008780:Methyltransferases / Q000302:isolation & purification*
D004247:DNA; D008780:Methyltransferases; D004248:DNA (Cytosine-5-)-Methyltransferases; D004720:Endonucleases; D004262:DNA Restriction Enzymes
Fraktsionirovanie i ochistka metilaz DNK i spetsificheskikh endonukleaz iz kletok Escherichia coli SK