Catalytic subunit of mitochondrial DNA polymerase from Drosophila embryos. Cloning, bacterial overexpression, and biochemical characterization

J Biol Chem. 1996 Sep 20;271(38):23389-94. doi: 10.1074/jbc.271.38.23389.


A full-length cDNA of the catalytic subunit of mitochondrial DNA polymerase from Drosophila embryos has been obtained, and its nucleotide sequence was determined. The cDNA clone encodes a polypeptide with a deduced amino acid sequence of 1145 residues and a predicted molecular mass of 129.9 kDa. Amino-terminal sequence analysis of the mature catalytic subunit of the heterodimeric mitochondrial enzyme from Drosophila embryos identified the amino-terminal amino acid at position +10 in the deduced amino acid sequence, indicating a mitochondrial presequence peptide of only nine amino acids. Alignment of the catalytic subunit sequence with that of Escherichia coli DNA polymerase I Klenow fragment indicated a high degree of amino acid sequence conservation in each of the three DNA polymerase and three 3' --> 5' exonuclease domains identified by biochemical studies in the latter enzyme. Bacterial overexpression, purification, and biochemical analysis demonstrated both 5' --> 3' DNA polymerase and 3' --> 5' exonuclease in the recombinant polypeptide. This represents the first demonstration of 3' --> 5' exonuclease activity in the polymerase catalytic subunit of animal mitochondrial DNA polymerase.

Publication types

  • Comparative Study
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • Cloning, Molecular
  • DNA Polymerase I / genetics
  • DNA, Complementary / genetics
  • DNA-Binding Proteins / metabolism
  • DNA-Directed DNA Polymerase / genetics*
  • DNA-Directed DNA Polymerase / metabolism*
  • Drosophila / embryology
  • Drosophila / enzymology
  • Escherichia coli / genetics
  • Exodeoxyribonuclease V
  • Exodeoxyribonucleases / metabolism*
  • Mitochondria / enzymology*
  • Mitochondria / genetics*
  • Molecular Sequence Data
  • Protein Binding
  • Protein Conformation
  • Recombinant Proteins / metabolism
  • Sequence Analysis, DNA
  • Sequence Homology, Amino Acid
  • Structure-Activity Relationship


  • DNA, Complementary
  • DNA-Binding Proteins
  • Recombinant Proteins
  • DNA Polymerase I
  • DNA-Directed DNA Polymerase
  • Exodeoxyribonucleases
  • Exodeoxyribonuclease V

Associated data

  • GENBANK/U62547