4FS2

Base pairing mechanism of N2,3-ethenoguanine with dCTP by human polymerase iota

Summary for 4FS2

• Entry DOI: 10.2210/pdb4fs2/pdb
• Related: 3OSN 3V6H 3V6J 3V6K 4FS1
• Descriptor: DNA (5'-D(*TP*CP*TP*(EFG)P*GP*GP*GP*TP*CP*CP*TP*AP*GP*GP*AP*CP*CP*(DOC))-3'), DNA polymerase iota, MAGNESIUM ION, ... (6 entities in total)
• Functional Keywords: human dna polymerase iota, dna polymerase, dna replication, lesion bypass, 2'-fluoro arabinose, n2, 3-ethenoguanine, dttp, template, transferase-dna complex, transferase/dna
• Biological source: Homo sapiens (human); More
• Cellular location: Nucleus: Q9UNA4
• Total number of polymer chains: 3
• Total formula weight: 58659.54

Authors

Zhao, L. (deposition date: 2012-06-26, release date: 2012-08-29, Last modification date: 2024-02-28)

Primary citation

Zhao, L.,Pence, M.G.,Christov, P.P.,Wawrzak, Z.,Choi, J.Y.,Rizzo, C.J.,Egli, M.,Guengerich, F.P.
Basis of Miscoding of the DNA Adduct N2,3-Ethenoguanine by Human Y-family DNA Polymerases.
J.Biol.Chem., 287:35516-35526, 2012

PubMed Abstract: N(2),3-Ethenoguanine (N(2),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-N(2),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 N(2),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 N(2),3-εG paired to dCTP and dTTP revealed Hoogsteen-like base pairing mechanisms. Two hydrogen bonds were observed in the N(2),3-εG:dCTP base pair, whereas only one appears to be present in the case of the N(2),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 N(2),3-εG.

PubMed: 22910910
DOI: 10.1074/jbc.M112.403253

Experimental method

X-RAY DIFFRACTION (2.05 Å)