Major Groove Orientation of the (2S)-N6-(2-Hydroxy-3-buten-1-yl)-2′-deoxyadenosine DNA Adduct Induced by 1,2-Epoxy-3-butene

1,3-Butadiene (BD) is an environmental and occupational toxicant classified as a human carcinogen. It is oxidized by cytochrome P450 monooxygenases to 1,2-epoxy-3-butene (EB), which alkylates DNA. BD exposures lead to large numbers of mutations at A:T base pairs even though alkylation of guanines is more prevalent, suggesting that one or more adenine adducts of BD play a role in BD mediated genotwdcity. However, the etiology of ED mediated genotoxicity at adenine remains poorly understood. EB alkylates the N-6 exocyclic nitrogen of adenine to form N-6-(hydroxy-3-buten-1-yl)-2'-dA ((2S)-N-6-HB-dA) adducts (Tretyakova, N., Lin, Y, Sangaiah, R, Upton, P. B, and Swenberg J. A. (1997) Carcinogenesis 18, 137-147). The structure of the (2S)-N-6-HB-dA adduct has been determined in the 5'-d(C(1)G(2)G(3)A(4)C(5)Y(6)G(8)A(9)A(10)G(11))-3':5'-d(C(12)T(13)T(14)C(15)T(16)T(17)G(18)T(19) C(20)C(21)G(22))-3' duplex [Y = (2S)-N-6-HB-dA] containing codon 61 (underlined) of the human N-ras protooncogene, from NMR spectroscopy. The (2S)-N6-HB-dA adduct was positioned in the major groove, such that the butadiene moiety was oriented in the 3 direction. At the C-alpha carbon, the methylene protons of the modified nudeobase Y-6 faced the 5' direction, which placed the C-beta carbon in the 3' direction. The C-beta hydroxyl group faced toward the solvent, as did carbons C-gamma and C-delta. The C-beta hydroxyl group did not form hydrogen bonds with either T-16 O-4 or T-17 O-4. The (2S)-N-6-HB-dA nudeoside maintained the anti conformation about the glycosyl bond, and the modified base retained Watson Crick base pairing with the complementary base (T-17). The adduct perturbed stacking interactions at base pairs C-5:G(18), Y-6:T-17, and A(7):T-16 such that the Y-6 base did not stack with its 5' and 3' neighbor C-5, but it did with its 3' neighbor A(7). The complementary thymine 747 stacked well with both 5' and 3' neighbors T16 and G18. The presence of the (2S)-N-6-FIB-dA resulted in a 5 degrees C reduction in the T-m of the duplex, which is attributed to less favorable stacking interactions and adduct accommodation in the major groove.