Isolation and characterization of BsuE methyltransferase, a CGCG specific DNA methyltransferase from Bacillus subtilis.

摘要: The DNA methyltransferase M-BsuE that recognizes the sequence 5'-CGCG-3' has been isolated from Bacillus subtilis strain ISE15. A 1600-fold purification of was achieved by column chromatography on phosphocellulose, heparin-Sepharose, and DEAE-Sepharose. activity monitored in eluants radiochemically transfer tritiated methyl groups radiolabeled S-adenosylmethionine to poly(dGdC)-poly(dGdC) DNA, a sensitive specific substrate for activity. specificity this confirmed enzymatically demonstrating M-BsuE-methylated selectively protected cleavage restriction enzyme isoschizomers, ThaI FnuDII. Purified an apparent molecular size 41,000-43,000 as determined gel filtration migrates 41-kDa protein sodium dodecyl sulfate-polyacrylamide gel. methylation is dependent upon presence 2-mercaptoethanol. optimal at 37 degrees C 50 mM Tris-HCl, pH 7.8, 25 KCl, 6 microM S-adenosylmethionine, 5 2-mercaptoethanol, 10 EDTA. methylates external cytidine its recognition both linear supercoiled DNA. unique property ability methylate Z-DNA.