Indistinguishable physical and catalytic properties of DNA methyltransferase from normal rat liver and a transplantable rat hepatocellular carcinoma.
DNA methyltransferase (DMase) was purified 700- and 1002-fold from normal rat liver and transplantable hepatocellular carcinoma 252 (THC 252) nuclei, respectively, using a four-step procedure that included chromatography on phosphocellulose, hydroxylapatite, DEAE-Sephacel and gel filtration on AcA 34. The enzymes had identical characteristics: pI = 7.4-7.6; Mr = 280 000 by gel filtration; preference for methylating double-stranded over single-stranded DNA and hemimethylated over unmethylated DNA templates; and apparent km of 10 microM for dinucleotide units in poly(dC-dG) and 0.5 microM for S-adenosylmethionine (SAM). Thermal inactivation profiles and sulfhydryl group alkylation inhibition curves for fraction III produced very similar single-transition curves, suggesting the presence of a single-functional enzyme species that is indistinguishable between normal and tumor tissue. Single-value Michaelis-Menten kinetics were obtained for fraction IV enzymes with respect to the concentration of SAM and dinucleotide units in poly(dC-dG), suggesting the absence of isozymic or multiple forms of DMase in normal and malignant liver tissues.
D000818:Animals; D004247:DNA; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000302:isolation & purification; D004277:DNA, Single-Stranded; D006358:Hot Temperature; D007526:Isoelectric Point; D008099:Liver / Q000201:enzymology*; D008114:Liver Neoplasms, Experimental / Q000201:enzymology*; D008780:Methyltransferases / Q000302:isolation & purification; D008970:Molecular Weight; D009368:Neoplasm Transplantation; D051381:Rats; D011912:Rats, Inbred ACI
D004277:DNA, Single-Stranded; D004247:DNA; D008780:Methyltransferases; D004248:DNA (Cytosine-5-)-Methyltransferases