Structure of a site specific major groove (2S,3S)-N⁶-(2,3,4- trihydroxybutyl)-2′-deoxyadenosyl DNA adduct of butadiene diol epoxide

The solution structure of the (2S,3S)-N⁶-(2,3,4-trihydroxybutyl) -2′-deoxyadenosyl adduct arising from the alkylation of adenine N ⁶ at position X⁶ in d(CGGACXAGAAG)·d(CTTCTTGTCCG), by butadiene diol epoxide, was determined. This oligodeoxynucleotide contains codon 61 (underlined) of the human N-ras protooncogene. This oligodeoxynucleotide, containing the adenine N⁶ adduct butadiene triol (BDT) adduct at the second position of codon 61, was named the ras61 S,S-BDT-(61,2) adduct. NMR spectroscopy revealed modest structural perturbations localized to the site of adduction at X⁶·T¹⁷, and its nearest-neighbor base pairs C⁵·G¹⁸ and A ⁷·T¹⁶. All sequential NOE connectivities arising from DNA protons were observed. Torsion angle analysis from COSY data suggested that the deoxyribose sugar at X⁶ remained in the C2′-endo conformation. Molecular dynamics calculations using a simulated annealing protocol restrained by a total of 442 NOE-derived distances and J coupling-derived torsion angles refined structures in which the BDT moiety oriented in the major groove. Relaxation matrix analysis suggested hydrogen bonding between the hydroxyl group located at the β-carbon of the BDT moiety and the T¹⁷ O⁴ of the modified base pair X ⁶·T¹⁷. The minimal perturbation of DNA induced by this major groove adduct correlated with its facile bypass by three Escherichia coli DNA polymerases in vitro and its weak mutagenicity [Carmical, J. R., Nechev, L. V., Harris, C. M., Harris, T. M., and Lloyd, R. S. (2000) Environ. Mol. Mutagen. 35, 48-56]. Overall, the structure of this adduct is consistent with an emerging pattern in which major groove adenine N⁶ alkylation products of styrene and butadiene oxides that do not strongly perturb DNA structure are not strongly mutagenic.