Structural Consequences of Epimerization of Thymine Glycol Lesions in Duplex DNA: Implications for DNA Repair

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Thymine glycol (Tg), 5,6-dihydroxy-5,6-dihydrothymine, forms in DNA by reaction of thymine with reactive oxygen species. It exists as two diastereomeric pairs of epimers, the 5R cis, trans pair (5R,6S;5R,6R) and the 5S cis, trans pair (5S,6R; 5S,6S). The 5R pair is more abundant. At 30 °C, a 70%:30% cis:trans ratio of epimers is present in this sequence when SR-Tg is opposite dA. For the cis epimer Tg and A remain in the Watson-Crick alignment. The Tg N3H imine and A N 6 amine protons undergo increased solvent exchange. Stacking between Tg and the 3'-neighbor G•C base pair is disrupted. The solvent accessible surface and T 2 relaxation of Tg increases. Molecular dynamics calculations predict that the axial conformation of the Tg CH 3 group is favored; propeller twisting of the Tg•A pair and hydrogen bonding between Tg OH6 and the N7 atom of the 3'-neighbor guanine alleviate steric clash with the 5'-neighbor base pair. Tg also destabilizes the 5'-neighbor G•C base pair. Under these conditions, the human NER protein XPA binds to the 5R-Tg lesion comparably to the C8-dG acetylaminoflourene (AAF) adduct, whereas XPC/HR23B binding of the Tg lesion is superior than to the AAF adduct. In comparison, this lesion is processed by the Escherichia coli UvrA and UvrABC proteins less efficiently than the C8-dG AAF adduct. The destabilization of two base pairs by the cis epimer may facilitate flipping both base pairs from the helix, enabling XPC/HR23B binding prior to recruitment of XPA. When 5R-Tg pairs opposite dG in this sequence context only the cis epimer is observed. Tg assumes the wobble orientation and stacks below the 5'-neighbor dG, while the mismatched dG stacks below the 5'-neighbor dC. Stacking between Tg and the 3'-neighbor G•C base pair is disrupted. Differences in base excision repair of the Tg•G and Tg•A pairs by hNEIL1 may be related to the wobble orientation of the cis Tg epimer in the Tg•G pair, and the lack of hydrogen bonding between the Tg OH groups and the N7 atom of the 3'-neighbor dG. Hydrogen bonding between Tg 6 OH6→G 7 N7 in the Tg•A pair may increase the energetic barrier with regard to flipping of the Tg lesion into the active site pocket of the glycosylase, hindering repair.