ATP(GTP)-dependent conversion of MVM parvovirus single-stranded DNA to its replicative form by a purified 10 S species of mouse DNA polymerase α

Abstract

A species of DNA polymerase α that is active in the ATP(GTP)-dependent conversion of MVM parvovirus single-stranded linear DNA to the duplex replicative form has been purified 4300-fold from Ehrlich ascites mouse tumour cells. The single-stranded → replicative form activity is maintained throughout ammonium sulfate precipitation, DEAE-cellulose, phosphocellulose and hydroxyapatite column chromatography and glycerol gradient sedimentation. Polypeptides with M r = 230 000, 220 000, 183 000, 157 000, 125 000, 70 000, 65 000, 62 000, 57 000, 53 000 and 48 000 copurify with the single-stranded → replicative form activity, which sediments at approx. 10 S. The M r = 183 000, 157 000 and 125 000 polypeptides exhibit catalytic activity when assayed in situ following SDS-polyacrylamide gel electrophoresis. The 10 S form of DNA polymerase α is functionally distinguishable from an 8.4 S form of the enzyme obtained from the same cells on the basis of single-stranded → replicative form activity. The single-stranded → replicative form activity of the 10 S enzyme is stable at 22°C for up to 3 h, but exhibits a half life of only 5 min at 45°C.