Partial purification of the Escherichia coli K-12 mec+ deoxyribonucleic acid-cytosine methylase: in vitro methylation completely protects bacteriophage lambda deoxyribonucleic acid against cleavage by R-EcoRII.
A procedure is described for the partial purification of the deoxyribonucleic acid (DNA)-cytosine methylases controlled by the RII plasmid and by the Escherichia coli mec+ gene. The two enzymes exhibit similar but distinct chromatographic behavior on diethylaminoethyl-cellulose and phosphocellulose. Preliminary studies on the two methylases indicate that they are indistinguishable with respect to their Km for S-adenosylmethionine and their pH (in tris (hydroxymethyl)aminomethane buffer) and NaCl concentration optima. In vitro methylation of various phage lambda DNA substrates by the mec'r RII enzyme modifies the DNA to a form that is completely resistant to double-stranded cleavage by the RII restriction endonuclease (R-EcoRII). These results are consistent with our earlier proposal that the mec8ethylase recognizes RII host specificity sites.
D003090:Coliphages; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000302:isolation & purification* / Q000378:metabolism; D004262:DNA Restriction Enzymes / Q000378:metabolism*; D004279:DNA, Viral / Q000378:metabolism*; D004720:Endonucleases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D008745:Methylation; D008780:Methyltransferases / Q000302:isolation & purification*; D012965:Sodium Chloride / Q000494:pharmacology
D004279:DNA, Viral; D012965:Sodium Chloride; D008780:Methyltransferases; D004248:DNA (Cytosine-5-)-Methyltransferases; D004720:Endonucleases; D004262:DNA Restriction Enzymes