Basis of miscoding of the DNA adduct N2,3-ethenoguanine by human Y-family DNA polymerases.

ABSTRACT: N(2),3-Ethenoguanine (N(2),3-?G) is one of the exocyclic DNA adducts produced by endogenous processes (e.g. lipid peroxidation) and exposure to bioactivated vinyl monomers such as vinyl chloride, which is a known human carcinogen. Existing studies exploring the miscoding potential of this lesion are quite indirect because of the lability of the glycosidic bond. We utilized a 2'-fluoro isostere approach to stabilize this lesion and synthesized oligonucleotides containing 2'-fluoro-N(2),3-?-2'-deoxyarabinoguanosine to investigate the miscoding potential of N(2),3-?G by Y-family human DNA polymerases (pols). In primer extension assays, pol ? and pol ? replicated through N(2),3-?G, whereas pol ? and REV1 yielded only 1-base incorporation. Steady-state kinetics revealed that dCTP incorporation is preferred opposite N(2),3-?G with relative efficiencies in the order of pol ? > REV1 > pol ? ? pol ?, and dTTP misincorporation is the major miscoding event by all four Y-family human DNA pols. Pol ? had the highest dTTP misincorporation frequency (0.71) followed by pol ? (0.63). REV1 misincorporated dTTP and dGTP with much lower frequencies. Crystal structures of pol ? with N(2),3-?G paired to dCTP and dTTP revealed Hoogsteen-like base pairing mechanisms. Two hydrogen bonds were observed in the N(2),3-?G:dCTP base pair, whereas only one appears to be present in the case of the N(2),3-?G:dTTP pair. Base pairing mechanisms derived from the crystal structures explain the slightly favored dCTP insertion for pol ? in steady-state kinetic analysis. Taken together, these results provide a basis for the mutagenic potential of N(2),3-?G.

SUBMITTER: Zhao L 

PROVIDER: S-EPMC3471744 | biostudies-literature | 2012 Oct

REPOSITORIES: biostudies-literature