RT Journal Article
SR Electronic
T1 DNA Polymerase α-Primase Complexes from Carcinogen-treated Chinese Hamster Ovary Cells
JF Cancer Research
JO Cancer Res
FD American Association for Cancer Research
SP 6894
OP 6901
VO 50
IS 21
A1 Brucker, Angelika
A1 Loeb, Lawrence A.
A1 Thielmann, Heinz Walter
YR 1990
UL http://cancerres.aacrjournals.org/content/50/21/6894.abstract
AB In order to investigate whether carcinogens induce alterations of the DNA polymerase α-primase complex we compared the physicochemical and catalytic properties and the fidelity of DNA synthesis of DNA polymerase α-primase complexes from carcinogen-treated and untreated Chinese hamster ovary cells. Complexes were purified by ion exchange or by immunoaffinity chromatography and both DNA-polymerizing activities and those of ancillary enzymes, such as RNA primase and exonuclease, were examined. Further characterization of the complexes included determination of the relative molecular masses, sedimentation coefficients, and diffusion coefficients, and measurements of the Kms for deoxynucleotide triphosphates and DNA templates, which were identical for the preparations from both carcinogen-treated and untreated cells. The fidelity of DNA polymerase α-primase complexes measured by the φX174am3 reversion assay was also similar in carcinogen-treated and untreated cells. Thus, a carcinogen-mediated induction of a DNA polymerase α-primase complex with low fidelity was not observed within the detection limits of the φX174 assay. RNA primase was found to be an ancillary enzyme activity of the DNA polymerase α from both carcinogen-treated and untreated cells; however, the RNA primase:DNA polymerase α activity ratio was significantly higher in DNA polymerase α-primase complexes from carcinogen-treated cells. These complexes also exhibited an at least 3 times greater velocity of synthesis with supercoiled or unprimed single-stranded DNAs as templates. Since the binding sites of DNA polymerase α-primase complexes for deoxynucleotide triphosphates and DNA templates were shown to be identical before and after treatment of cells with carcinogens (i.e., identical Km values for different DNA templates and Ki values for specific inhibitors), the increased synthesis catalyzed by the DNA polymerase α-primase complex from carcinogen-treated cells might be due to a carcinogen-induced alteration of an accessory protein of the complex. ©1990 American Association for Cancer Research.