Abstract
Ionizing radiation induces oxidation of DNA precursor dNTPs in the nucleotide pool. The oxidized dNTPs can be incorporated into nascent DNA during DNA replication, resulting mutations and cell death. Previously, we have shown that incorporation of oxidized dNTP may be involved in Y-family polymerases. In particular, human polymerase η (hPolη) exhibited the activity to incorporation of 8oxodGTP comparable with normal dNTP. To elucidate the mechanism for incorporation of oxidized dNTP by hPolη, the enzymes replacing R61 were prepared and their activities were assessed. R61 might lie adjacent to the dNTP in hPolη structure inferred from the crystal structures of yeast Polη and DNA-Dpo4, an archaeal homologue of DinB. R61K exhibited the different specificity from wild-type for incorporation of oxidized dNTP; R61K elevated the activity to incorporated 8oxodGTP opposite dC and 8oxodATP opposite dT, although the specificity for incorporation of 2OHdATP was not different from wild-type. The activity bypassing 8oxodG or 8oxodA in the template strand did not differ between R61K and wild-type. These results indicated that R61 in hPolη is liable to induce the mutation by restricting to incorporate C8-oxidized dNTP.
