Interconversion of the cis-5R,6S- and trans-5R,6R-Thymine Glycol Lesions in Duplex DNA

Kyle L. Brown†, Travis Adams†, Vijay P. Jasti‡, Ashis K. Basu‡ and Michael P. Stone*†
Department of Chemistry, Center in Molecular Toxicology, Vanderbilt-Ingram Cancer Center, Vanderbilt University, Nashville, Tennessee 37235, and Department of Chemistry, University of Connecticut, Storrs, Connecticut 06269
J. Am. Chem. Soc., 2008, 130 (35), pp 11701–11710
DOI: 10.1021/ja8016544
Publication Date (Web): August 6, 2008
Copyright © 2008 American Chemical Society
†

Vanderbilt University.

,
‡

University of Connecticut.

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Abstract

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Thymine glycol (Tg), 5,6-dihydroxy-5,6-dihydrothymine, is formed in DNA by the reaction of thymine with reactive oxygen species. The 5R Tg lesion was incorporated site-specifically into 5′-d(G1T2G3C4G5Tg6G7T8T9T10G11T12)-3′; Tg = 5R Tg. The Tg-modified oligodeoxynucleotide was annealed with either 5′-d(A13C14A15A16A17C18A19C20G21C22A23C24)-3′, forming the Tg6•A19 base pair, corresponding to the oxidative damage of thymine in DNA, or 5′-d(A13C14A15A16A17C18G19C20G21C22A23C24)-3′, forming the mismatched Tg6•G19 base pair, corresponding to the formation of Tg following oxidative damage and deamination of 5-methylcytosine in DNA. At 30 °C, the equilibrium ratio of cis-5R,6S:trans-5R,6R epimers was 7:3 for the duplex containing the Tg6•A19 base pair. In contrast, for the duplex containing the Tg6•G19 base pair, the cis-5R,6S:trans-5R,6R equilibrium favored the cis-5R,6S epimer; the level of the trans-5R,6R epimer remained below the level of detection by NMR. The data suggested that Tg disrupted hydrogen bonding interactions, either when placed opposite to A19 or G19. Thermodynamic measurements indicated a 13 °C reduction of Tm regardless of whether Tg was placed opposite dG or dA in the complementary strand. Although both pairings increased the free energy of melting by 3 kcal/mol, the melting of the Tg•G pair was more enthalpically favored than was the melting of the Tg•A pair. The observation that the position of the equilibrium between the cis-5R,6S and trans-5R,6R thymine glycol epimers in duplex DNA was affected by the identity of the complementary base extends upon observations that this equilibrium modulates the base excision repair of Tg [Ocampo-Hafalla, M. T.; Altamirano, A.; Basu, A. K.; Chan, M. K.; Ocampo, J. E.; Cummings, A., Jr.; Boorstein, R. J.; Cunningham, R. P.; Teebor, G. W.DNA Repair (Amst)2006, 5, 444−454].

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History

  • Published In Issue September 03, 2008
  • Article ASAPAugust 06, 2008
  • Received: March 5, 2008

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