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(<i>R</i>)-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(CGGA<u>CXA</u>GAAG)-3‘·5‘-d(CTTCTTGTCCG)-3‘. This oligodeoxynucleotide contained the BDO adduct at the second position of codon 61 of the human <i>N-ras</i> protooncogene (underlined) and was named the <i>ras61</i> <i>R</i>-N1-BDO-(61,2) adduct. <sup>1</sup>H NMR revealed a weak C<sup>5</sup> H1‘ to X<sup>6</sup> H8 nuclear Overhauser effects (NOE), followed by an intense X<sup>6</sup> H8 to X<sup>6</sup> H1‘ NOE. Simultaneously, the X<sup>6</sup> H8 to X<sup>6</sup> H3‘ NOE was weak. The resonances arising from the T<sup>16</sup> and T<sup>17</sup> imino protons were not observed. <sup>1</sup>H 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 T<sup>17</sup>, 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 <i>Escherichia coli</i> [Rodriguez, D. A., Kowalczyk, A., Ward, J. B. J., Harris, C. M., Harris, T. M., and Lloyd, R. S. (2001) <i>Environ. Mol. Mutagen. 38</i>, 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) <i>Chem. Res. Toxicol. 15</i>, 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 <i>R</i>- and the <i>S</i>-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.