PRIMARY STRUCTURE OF THE CATALYTIC SUBUNIT OF CALF THYMUS DNA POLYMERASE-DELTA - SEQUENCE SIMILARITIES WITH OTHER DNA-POLYMERASES

The 125- and 48-kDa subunits of bovine DNA polymerase-delta have been isolated by SDS-polyacrylamide gel electrophoresis and demonstrated to be unrelated by partial peptide mapping with N-chlorosuccinimide. A 116-kDa polypeptide, usually present in DNA polymerase-delta preparations, was shown to be a degraded form of the 125-kDa catalytic subunit. Amino acid sequence data from Staphylococcus aureus V8 protease, cyanogen bromide, and trypsin digestion of the 125- and 116-kDa polypeptides were used to design primers for the polymerase chain reaction to determine the nucleotide sequence of a full-length cDNA encoding the catalytic subunit of bovine DNA polymerase-delta. The predicted polypeptide is 1106 amino acids in length with a calculated molecular weight of 123 707. This is in agreement with the molecular weight of 125 000 estimated from SDS-polyacrylamide gel electrophoresis. Comparison of the deduced amino acid sequence of the catalytic subunit of bovine DNA polymerase-delta with that of its counterpart from Saccharomyces cerevisiae showed that the proteins are 44% identical. The catalytic subunit of bovine DNA polymerase-delta contains the seven conserved regions found in a number of bacterial, viral, and eukaryotic DNA polymerases. It also contains five additional regions that are highly conserved between bovine and yeast DNA polymerase-delta, but these regions share little or no homology with the alpha-polymerases. Four of these additional regions are also highly homologous to the herpes virus family of DNA polymerases, whereas one region is not homologous to any other DNA polymerase that has been sequenced thus far. The polypeptide also contains two C-terminal clusters of cysteine residues postulated to be DNA binding sites or zinc ringers.