Temperature-dependence of UV Laser One-electron Oxidative Guanine Modifications as a Probe of Local Stacking Fluctuations and Conformational Transitions

By monitoring Rpip/RFpg, i.e. the relative sensitivity to hot piperidine and to formamidopyrimidine DNA glycosylase (Fpg protein) of the guanine lesions induced in DNA exposed to UV laser irradiation, we have previously observed that the formation of the two major types of one-electron oxidative guanine modifications, oxazolone and 7,8-dihydro-8-oxoguanine (8-oxodG), depends on DNA conformational features. While oxazolone is largely predominant at each site of single-stranded DNA (Rpip>RFpg), 8-oxodG is the major lesion at most of the sites of double-stranded DNA (RpipRFpg at 20 °C and the ratio Rpip/RFpg does not vary significantly during the melting process. Interestingly, these guanine residues display a high sensitivity to dimethyl sulfoxide methylation while the opposite cytosine residues are unsensitive, suggesting that the prevalence of Rpip over RFpg is related not to base-pairing disruption but rather to the local helical alteration of the B-DNA stacking geometry. This leads us to propose that the slight variations in the ratios Rpip/RFpg observed, at individual sites, at temperatures below the helix–coil transition reflect local small-scale breathing motions, unstacking single dinucleotide steps prior to opening. Our results thus support the view that the temperature dependence of the ratio of Rpip/RFpg at sites of B-DNA provides a sensitive probe of the DNA internal local thermal stability and are discussed in relation with the mechanisms proposed for the intramolecular rearrangement of the guanyl radical.