A multiprotein form of DNA polymerase α from HeLa cells. Resolution of its associated catalytic activities

Jamboor K. Vishwanatha, S. A. Coughlin, M. Wesolowski-Owen, E. F. Baril

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Abstract

The majority of the DNA polymerase α activity in HeLa cells has been isolated and purified as a multiprotein M(r) 640,000 form. The multiprotein form of DNA polymerase α corresponds to DNA polymerase α2 that was previously reported by us (Lamothe, P., Baril, B., Chi, A., Lee, L., and Baril, E. (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 4723-4727). The highly purified DNA polymerase α2 has in addition to DNA polymerase α-associated DNAse, primase, and diadenosine 5',5-P1,P4-tetraphosphate (Ap4A) binding activities and accessory primer recognition proteins C1 and C2. The DNA polymerase α and associated activities increase coordinately during the G1/S-phase transition of the cell cycle. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of the electrophoretically homogeneous DNA polymerase α shows that it is composed of at least eight polypeptides in the molecular weight range of 180,000-15,000. Hydrophobic chromatography on butyl-agarose resolves the DNAse and Ap4A-binding protein from a complex of DNA polymerase α, primase, and the primer recognition proteins C1 and C2. Hydrophobic chromatography of the latter complex on phenyl-Sepharose resolves the C1 protein from a DNA polymerase α-C2 protein-primase complex. Phosphocellulose chromatography of the DNA polymerase-primase-C2 protein complex resolves the C2 protein from a complex of DNA polymerase α-primase.

Original languageEnglish
Pages (from-to)6619-6628
Number of pages10
JournalJournal of Biological Chemistry
Volume261
Issue number14
Publication statusPublished - 1 Dec 1986

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