Identification of the oxidized products formed upon reaction of Chromium(V) with thymidine nucleotides

Kent D. Sugden, Karen E. Wetterhahn

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Two pathways have been observed for Cr(V)-mediated nucleotide oxidation in reactions of bis(2-ethyl-2-hydroxybutyrato)oxochromate(V) [CrO(ehba)2]- with thymidine nucleotides dTMP, dTDP, and dTTP. The extent of Cr(V)-induced nucleotide oxidation was greatest for thymidine diphosphate (dTDP), as measured by the production of thiobarbituric acid reactive species (TBARS; indicative of pathway 1) and thymine release (indicative of pathway 2). The nucleoside thymidine showed no reaction, suggesting a phosphate-dependent oxidation. Amounts of TBARS and thymine were maximal at a pH range of 6.0-6.5, and both TBARS formation and thymine release correlated with decay of the Cr(V) EPR signal. Formation of TBARS was maximal in 100% O2 but decreased markedly under argon, whereas thymine release was maximal under argon, but remained the major product observed under aerobic conditions. Pathway 1 for the reaction of Cr(V) with dTDP led to formation of glycolic acid and trans-thymine propenal at approximately equimolar amounts, consistent with a mechanism involving oxygen-dependent sugar oxidation following hydrogen atom abstraction at the C-4' carbon of the deoxyribose sugar. Pathway 2 led to release of free thymine, but much less (barely detectable) 2-deoxy-D-pentitol was formed from postreduction of the reactive aldehydic sugar fragment. Thus, the oxygen-independent release of thymine does not appear to result from reaction at the C-4' hydrogen unless decomposition of the aldehydic intermediate occurred. Determination of the oxidation state of chromium responsible for the observed oxidative damage was carried out using Mn(II), a Cr(IV)-specific reductant. Mn(II) essentially abolished all activity for both TBARS formation (pathway 1) and thymine release (pathway 2). These results suggest that Cr(IV), formed upon disproportionation of Cr(V), oxidizes the nucleotide deoxyribose sugar moiety via a phosphate-bound intermediate. pathway 1 involves oxygen-dependent oxidation at the C-4' position; however, the mechanism for oxygen-independent thymine release (pathway 2) is still unclear.

Original languageEnglish
Pages (from-to)10811-10818
Number of pages8
JournalJournal of the American Chemical Society
Issue number44
StatePublished - Nov 6 1996


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