8-oxodGTP incorporation by DNA polymerase beta is modified by active-site residue Asn279

Biochemistry. 2000 Feb 8;39(5):1029-33. doi: 10.1021/bi991789x.


To understand how the active site of a DNA polymerase might modulate the coding of 8-oxo-7,8-dihydrodeoxyguanine (8-oxodG), we performed steady-state kinetic analyses using wild-type DNA polymerase beta (pol beta) and two active-site mutants. We compared the coding of these polymerases by calculating the ratio of efficiencies for incorporation of dATP and dCTP opposite 8-oxodG and for incorporation of 8-oxodGTP opposite dA and dC. For wild-type pol beta, there is a 2:1 preference for incorporation of dCTP over dATP opposite 8-oxodG using a 5'-phosphorylated 4-base gap substrate. Mutation of either Asn279 or Arg283 to alanine has almost no effect on the ratio. 8-OxodGTP is preferentially incorporated opposite a template dA (24:1) by wild-type pol beta; mutation of Asn279 to alanine results dramatic change whereby there is preferential incorporation of 8-oxodGTP opposite dC (14:1). This suggests that interactions of 8-oxodGTP with Asn279 in the polymerase active site may alter the conformation of 8-oxodGTP and therefore alter its misincorporation.

Publication types

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

MeSH terms

  • 8-Hydroxy-2'-Deoxyguanosine
  • Asparagine / genetics
  • Asparagine / metabolism*
  • Binding Sites / genetics
  • DNA Polymerase beta / genetics
  • DNA Polymerase beta / metabolism*
  • Deoxyguanine Nucleotides / genetics
  • Deoxyguanine Nucleotides / metabolism*
  • Deoxyguanosine / analogs & derivatives
  • Deoxyguanosine / metabolism
  • Humans
  • Kinetics
  • Mutagens / metabolism*
  • Nucleic Acid Conformation
  • Oligodeoxyribonucleotides / chemical synthesis
  • Oligodeoxyribonucleotides / metabolism
  • Substrate Specificity / genetics
  • Templates, Genetic


  • Deoxyguanine Nucleotides
  • Mutagens
  • Oligodeoxyribonucleotides
  • 8-oxodeoxyguanosine triphosphate
  • Asparagine
  • 8-Hydroxy-2'-Deoxyguanosine
  • DNA Polymerase beta
  • Deoxyguanosine