Six Germline Genetic Variations Impair the Translesion Synthesis Activity of Human DNA Polymerase κ.

DNA polymerase (pol) κ efficiently catalyzes error-free translesion DNA synthesis (TLS) opposite bulky N-guanyl lesions induced by carcinogens such as polycyclic aromatic hydrocarbons. We investigated the biochemical effects of nine human nonsynonymous germline POLK variations on the TLS properties of pol κ, utilizing recombinant pol κ (residues 1-526) enzymes and DNA templates containing an N-CH(9-anthracenyl)G (N-AnthG), 8-oxo-7,8-dihydroguanine (8-oxoG), O-methyl(Me)G, or an abasic site. In steady-state kinetic analyses, the R246X, R298H, T473A, and R512W variants displayed 7- to 18-fold decreases in k/K for dCTP insertion opposite G and N-AnthG, with 2- to 3-fold decreases in DNA binding affinity, compared to that of the wild-type, and further showed 5- to 190-fold decreases in k/K for next-base extension from C paired with N-AnthG. The A471V variant showed 2- to 4-fold decreases in k/K for correct nucleotide insertion opposite and beyond G (or N-AnthG) compared to that of the wild-type. These five hypoactive variants also showed similar patterns of attenuation of TLS activity opposite 8-oxoG, O-MeG, and abasic lesions. By contrast, the T44M variant exhibited 7- to 11-fold decreases in k/K for dCTP insertion opposite N-AnthG and O-MeG (as well as for dATP insertion opposite an abasic site) but not opposite both G and 8-oxoG, nor beyond N-AnthG, compared to that of the wild-type. These results suggest that the R246X, R298H, T473A, R512W, and A471V variants cause a general catalytic impairment of pol κ opposite G and all four lesions, whereas the T44M variant induces opposite lesion-dependent catalytic impairment, i.e., only opposite O-MeG, abasic, and bulky N-G lesions but not opposite G and 8-oxoG, in pol κ, which might indicate that these hypoactive pol κ variants are genetic factors in modifying individual susceptibility to genotoxic carcinogens in certain subsets of populations.