Characterization of dna polymerase α from untransformed and pSV3.neo-transformed human fibroblasts

Abstract

1. 1. The specific activity of DNA-polymerase α isolated from pSV3.neo-transformed cells was more than 9-fold higher than that of polymerase a from untransformed cells. 2. 2. Western blot analysis, using anti-SV40 large T antigen, of both a crude cellular extract and of partially purified polymerase a from pSV3.neo-transformed cells revealed a single 76 kDa immunoreactive band not found in either crude extracts or partially purified enzyme from untransformed cells. 3. 3. The α polymerases from untransformed and transformed cells differed in molecular size, sensitivity to various inhibitors, specificity of template-primer utilization, and binding affinity for DNA cellulose, but showed essentially no differences in K m or V max . 4. 4. These data suggest that polymerase α isolated from pSV3.neo-transformed cells exhibits altered physical and catalytic characteristics compared with its untransformed cell counterpart, and that those alterations may be associated with increased replication of the genome in plasmid-transformed cells.