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 3...

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Bibliographic Details
Published in:Carcinogenesis (New York) 1985-06, Vol.6 (6), p.877-882
Main Authors: Ruchirawat, Mathuros, Becker, Frederick F., Lapeyre, Jean-Numa
Format: Article
Language:English
Subjects:
Animals
Biological and medical sciences
DNA
DNA (Cytosine-5-)-Methyltransferases - isolation & purification
DNA, Single-Stranded
General aspects (metabolism, cell proliferation, established cell line...)
Hot Temperature
Isoelectric Point
Liver - enzymology
Liver Neoplasms, Experimental - enzymology
Medical sciences
Methyltransferases - isolation & purification
Molecular Weight
Neoplasm Transplantation
Rats
Rats, Inbred ACI
Tumor cell
Tumors
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Summary: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 singlestranded DNA and hemimethylated over unmethylated DNA templates; and apparent km of 10 μM for dinucleotide units in poly(dC-dG) and 0.5 μM 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.
ISSN:0143-3334
1460-2180
DOI:10.1093/carcin/6.6.877