8-OxodGTP incorporation by DNA polymerase β is modified by active-site residue Asn279

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 β (pol β) 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 β, 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 β; 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.