Structural and conformational analyses of 8-hydroxy-2'-deoxyguanosine.

Oxidative DNA damage has been shown to involve formation of 8-hydroxy-2'-deoxyguanosine, which may serve as a mispairing lesion during cellular DNA replication. In order to assess the mutagenic potential of this DNA adduct, we examined the possible occurrence of several tautomeric forms and of different base conformations about the deoxyribose. Several spectroscopic and electronic absorption techniques were employed and showed structural changes occurring over a broad pH range. Two pKa's near pH 8 and 12 were observed by pH-solvent partitioning experiments, ultraviolet absorption spectral analyses, and 13C NMR spectroscopic methods. The presence of two pKa's suggested the formation of a dianion, with the second being formed in strong alkali. A comparison of ultraviolet absorption band features of 8-hydroxy-2'-deoxyguanosine with that of different C6,C8-diketo or enol derivatives supported a C8-keto assignment and also provided evidence that this DNA adduct contains a C6-keto group at physiological pH. 13C NMR data showed marked chemical shifts at C6 in solutions of pH 7.4-9.3, indicating the location of the first ionization. Increasing basicity produced further shifts at C5 and C8, indicating the C8 position for the second ionization. Multiple infrared bands were observed in the carbonyl region of the neutral compound, but only a single carbonyl band at 1692 cm-1 remained at pH 9.1, implying a keto group at C8. Determination of the chemical shifts and the nuclear Overhauser enhancements of the N1 and N7 resonances in the proton-decoupled 15N NMR spectrum indicated that both nitrogens were indeed protonated at neutral pH.(ABSTRACT TRUNCATED AT 250 WORDS)