DNA-cytosine-5-methyltransferase from P815 mouse mastocytoma cells: “Maintenance” and “de novo” activities are carried out by the same enzyme molecule

Abstract 1. 1. DNA-5-methyltransferase has been purified (about 1400-fold) from rapidly proliferating mouse P815 mastocytoma cells by chromatographies on DEAE cellulose, hydroxyapatite and a heparineagarose affinity step. 2. 2. The isolated enzyme has an isoelectric point of 7.3 and in neutral 10–30% glycerol gradient it bands in an area corresponding to molecular weight of 135,000 dalton. 3. 3. During the enzymatic reaction, the enzyme first interacts with DNA and then accomplishes a series of methyl group transfers without being detached. 4. 4. The formation of the initial DNA-enzyme complexes is probably random and independent of the cofactor, S- adenosyl- l -methionine , as well as the sequences recognized as methylation sites. 5. 5. The “maintenance” and “de novo”types of activity have been monitored using hemimethylated and completely unmethylated DNA as methyl group accepting polymers. 6. 6. Both these activities copurify in three different Chromatographie procedures. 7. 7. This, together with the fact that the enzyme purified near to homogeneity possesses both types of activities suggests that “de novot and “maintenance” DNA methyltransferase activities are exercised by the same enzyme molecule.