PROTONATION OF NUCLEOBASE ANIONS IN GAMMA-IRRADIATED DNA AND MODEL SYSTEMS - WHICH DNA-BASE IS THE ULTIMATE SINK FOR THE ELECTRON

Electron spin resonance (ESR) spectroscopy has been used to investigat e irreversible protonations of the nucleobase anions in gamma-irradiat ed frozen aqueous solutions of dGMP.dCMP, polyG.polyC, poly[dGdC] poly [dGdC], dAMP.dTMP, poly[dAdT].poly[dAdT] and DNA itself. Analysis of t he ESR spectra at a dose of 22 kGy shows that fractional conversion of total radicals to carbon-protonated species on annealing is in the or der: dAMP.dTMP (43%) > pdAdT = DNA (23%) > dGMP.dCMP (15%) > polydGdC. polydGdC (6%) 1 polyG.polyC (3%). Two hydrogen addition radicals make contributions to the polyG.polyC, poly[dGdC].poly[dGdC] and dGMP.dCMP spectra in H2O on annealing. They are those formed by protonations at C6 of the cytosine anion radical, C(C6)H-., and at C8 of the guanine a nion radical, G(C6)H-.. Computer analysis reveals that anion protonati on reaction in dGMP.dGMP results in mainly C(C6)H-., whereas protonati on reaction in polyG.polyC and poly[dGdC].poly[dGdC] yields mainly G(C 8)H-.. In dAMP.dTMP and poly[dAdT].poly[dAdT] as in DNA itself, the on ly DNA base found to undergo an irreversible protonation at a carbon s ite is thymine, resulting in T(C6)H-.. The conversion of DNA anion to T(C6)H-. is found to be dependent on dose. At low doses (5 kGy), about 30% conversion to T(C6)H-. is found, whereas at high doses (94 kGy), only 13% conversion is found. The dose dependence is ascribed in part to ion radical recombinations whose probabilities are increased at hig h doses. A consideration of the rates of protonations of the purine an d pyrimidine anion radicals as well as the differences in electron aff inities suggest carbon protonation reactions of DNA base anions in irr adiated stacked double-strand DNA at 37 degrees C would be predominant ly at thymine and perhaps guanine, whereas in single-strand DNA all ba ses would contribute.