Dual roles of glycosyl torsion angle conformation and stereochemical configuration in butadiene oxide-derived N1 β-hydroxyalkyl deoxyinosine adducts: A structural perspective

The solution structure of the N1-[1-hydroxy-3-buten-2(R)-yl]-2′- deoxyinosine adduct arising from the alkylation of adenine N1 by butadiene epoxide (BDO), followed by deamination to deoxyinosine, was determined in the oligodeoxynucleotide 5′-d(CGGACXAGAAG)-3′-5′-d(CTTCTTGTCCG)- 3′. This oligodeoxynucleotide contained the BDO adduct at the second position of codon 61 of the human N-ras protooncogene (underlined) and was named the ras61 R-N1-BDO-(61,2) adduct. ¹H NMR revealed a weak C ⁵ H1′ to X⁶ H8 nuclear Overhauser effects (NOE), followed by an intense X⁶ H8 to X⁶ H1′ NOE. Simultaneously, the X⁶ H8 to X⁶ H3′ NOE was weak. The resonances arising from the T¹⁶ and T¹⁷ imino protons were not observed. ¹H NOEs between the butadiene moiety and the DNA positioned the adduct in the major groove. Structural refinement based upon a total of 394 NOE-derived distance restraints and 151 torsion angle restraints yielded a structure in which the modified deoxyinosine was in the syn conformation about the glycosyl bond, with a glycosyl bond angle of 83°, and T¹⁷, the complementary nucleotide, was stacked into the helix but not hydrogen bonded with the adducted inosine. The refined structure provides a plausible hypothesis as to why these N1 deoxyinosine adducts strongly code for the incorporation of dCTP during trans lesion DNA replication, irrespective of stereochemistry, both in Escherichia coli [Rodriguez, D. A., Kowalczyk, A., Ward, J. B. J., Harris, C. M., Harris, T. M., and Lloyd, R. S. (2001) Environ. Mol. Mutagen. 38, 292-296] and in mammalian cells [Kanuri, M., Nechev, L. N., Tamura, P. J., Harris, C. M., Harris, T. M., and Lloyd, R. S. (2002) Chem. Res. Toxicol. 15, 1572-1580]. Rotation of the N1 deoxyinosine adduct into the syn conformation may facilitate incorporation of dCTP via Hoogsteen type templating with deoxyinosine, generating A to G mutations. However, conformational differences between the R- and the S-N1-BDO-(61,2) adducts, involving the positioning of the butenyl moiety in the major groove of DNA, suggest that adduct stereochemistry plays a secondary role in modulating the biological response to these adducts.