Mouse DNA polymerase alpha. Subunit structure and identification of a species with associated exonuclease.
Two species of alpha-polymerase with very similar catalytic properties have been purified to near homogeneity from a soluble protein fraction of mouse myeloma. Sedimentation analysis in 0.5 M salt-containing glycerol gradients indicated that both species had a native Mr of about 190,000. Each species contained nonidentical subunits with apparent molecular weights of about 47,000 and 54,000. Subunits of Mr = approximately 50,000 had been found previously in calf thymus alpha-polymerase (Holmes, A. M., Hesslewood, I. P., and Johnston, I. R. (1974) Eur. J. Biochem. 43, 487-499; (1976) Eur. J. Biochem. 62, 229-235). Tryptic peptide mapping failed to reveal primary structure homology between the subunits of the two enzymes. Thus, the two alpha-polymerases are clearly different species. These two enzymes are further distinguished by the fact that one of them has associated exonuclease activities. One activity degraded single-stranded DNA to mononucleotides in the 3' leads to 5' direction and acted distributively. The other exonuclease activity also degraded single-stranded DNA to mononucleotides, but this degradation was in the 5' leads to 3' direction in a processive fashion. Both exonuclease activities co-migrated with the polymerase activity during the final purification step of polyacrylamide gradient gel electrophoresis, which yielded the essentially homogenous alpha-polymerase, and also during sedimentation of the purified enzyme through a high salt glycerol gradient.
D000818:Animals; D002460:Cell Line; D004257:DNA Polymerase II / Q000302:isolation & purification / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D005092:Exonucleases / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D051379:Mice; D008970:Molecular Weight; D010954:Plasmacytoma; D013379:Substrate Specificity; D013698:Templates, Genetic
D046911:Macromolecular Substances; D004257:DNA Polymerase II; D004259:DNA-Directed DNA Polymerase; D005092:Exonucleases