Mutagenesis of a highly conserved lysine 340 of the PRD1 DNA polymerase

Biochim Biophys Acta. 1994 Oct 18;1219(2):260-6. doi: 10.1016/0167-4781(94)90047-7.


All known family B DNA polymerases contain a conserved region of amino acids, KX6-7YG, which appears to be correspond to the 'finger' alpha helix O of the Klenow fragment of E. coli DNA polymerase I, a family A DNA polymerase. Toward the goal of establishing the evolutionary relationship between the family A and B DNA polymerases, we have employed site-directed mutagenesis to access the functional role of the invariant amino acid lysine-340 of the PRD1 DNA polymerase. We have replaced the lysine-340 with three amino acids: histidine, asparagine and glutamic acid, respectively. Mutant DNA polymerases were overexpressed and purified to near homogeneity. Our results showed that the modification of the lysine-340 of the PRD1 DNA polymerase abolishes the polymerase activity without affecting the 3' to 5' exonuclease activity. These results support the proposal that the KX6-7YG motif of the family B DNA polymerases may be analogous to the KX7YG motif of the family A DNA polymerases, suggesting that two family DNA polymerases share a common ancestor.

Publication types

  • Comparative Study

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • Catalysis
  • Coliphages / enzymology*
  • DNA / biosynthesis
  • DNA Primers / chemistry
  • DNA Replication
  • DNA-Directed DNA Polymerase / chemistry*
  • Exonucleases / metabolism
  • Lysine / chemistry
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Sequence Alignment
  • Sequence Homology, Amino Acid
  • Structure-Activity Relationship


  • DNA Primers
  • DNA
  • DNA-Directed DNA Polymerase
  • Exonucleases
  • Lysine