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

Linlin Zhao, Matthew G. Pence, Plamen P. Christov, Zdzislaw Wawrzak, Jeong Yun Choi, Carmelo J. Rizzo, Martin Egli, F. Peter Guengerich*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

19 Scopus citations

Abstract

N2,3-Ethenoguanine (N2,3-εG) 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-N2,3-ε- 2′-deoxyarabinoguanosine to investigate the miscoding potential of N 2,3-εG by Y-family human DNA polymerases (pols). In primer extension assays, pol η and pol κ replicated through N 2,3-εG, whereas pol ι and REV1 yielded only 1-base incorporation. Steady-state kinetics revealed that dCTP incorporation is preferred opposite N2,3-εG with relative efficiencies in the order of pol κ > REV1 > pol η ≈ pol ι, and dTTP misincorporation is the major miscoding event by all four Y-family human DNA pols. Pol ι had the highest dTTP misincorporation frequency (0.71) followed by pol η (0.63). REV1 misincorporated dTTP and dGTP with much lower frequencies. Crystal structures of pol ι with N2,3-εG paired to dCTP and dTTP revealed Hoogsteen-like base pairing mechanisms. Two hydrogen bonds were observed in the N2,3-εG:dCTP base pair, whereas only one appears to be present in the case of the N2,3- εG:dTTP pair. Base pairing mechanisms derived from the crystal structures explain the slightly favored dCTP insertion for pol ι in steady-state kinetic analysis. Taken together, these results provide a basis for the mutagenic potential of N2,3-εG.

Original languageEnglish (US)
Pages (from-to)35516-35526
Number of pages11
JournalJournal of Biological Chemistry
Volume287
Issue number42
DOIs
StatePublished - Oct 12 2012

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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