Structural Studies of Fluxional Lesions in Deoxyribonucleic Acid

Abstract

This dissertation involves the structural and equilibrium studies of three DNA lesions: aflatoxin B1 formamidopyrimidine, methyl formamidopyrimidine, and thymine glycol. Studies of the methyl formamidopyrimidine (Me-dGuo-FAPY) found that the lesion exists as a mixture of alpha and beta anomers in the nucleotide 5'-CTATXATTCA-3'. Each anomer was in a 1:1 equilibrium and was a mix of rotamers. Aflatoxin B1 formamidopyrimidine adduct (AFB1-FAPY) was a mixture of alpha and beta anomers (2:1) in 5'-CTATXATTCA-3'; only the beta anomer was observed in 5'-CTATXATTCA-3'•5'-TGAATCATAG-3'. The alpha anomer destabilized duplex DNA by 13 °C while beta anomer stabilized the duplex 14 °C. Duplex stabilization of beta-AFB1-FAPY modified DNA was attributed in part to favorable stacking interactions with the complementary strand. Destabilization of alpha-AFB1-FAPY modified DNA was attributed to abnormal backbone geometry and disrupted stacking patterns with neighboring base pairs. The E geometrical isomer of the formamide was stabilized by an intra-strand hydrogen bond with the 3'-adenine N6 amino group; stabilization was reduced in single strand DNA so that the Z geometrical isomer was observed. In structural studies of AFB1-FAPY in oligonucleotides only the Ra atropisomer was observed. The alpha anomer of cis-5R,6S-thymine glycol lesion was not observed in single strand or duplex DNA. However, equilibrium of cis-5R,6S-thymine glycol with its trans epimer 5R,6R was 7:3 when base paired opposite adenine; only 5R,6S was observed when base paired with guanine. Thermodynamic analysis of cis-5R,6S-thymine glycol opposite adenine and guanine indicated that both lesions destabilize duplex DNA by 13 °C.