Effect of mismatched complementary strands and 5′-change in sequence context on the thermodynamics and structure of benzo[a]pyrene-modified oligonucleotides

Benzo[rr]pyrene (B[a]P) is a well-studied environmental carcinogen that when activated can react with DNA to form four major adducts: (+)-trans-, (-)-trans-, (+)-cis-, and (-)-cis-anti-B[a]P-dG. In this study, two oligonucleotides (5'-dCCATT-G(B[a]P)-CTACC-3' and 5'-dCCATC-G(B[a]P)-CTACC-3') were prepared, each containing the four isomeric adducts, and these were hybridized to either complementary sequences or to sequences containing an A, G, or T opposite the adducted guanine. Thermal melting curves, CD, and UV spectra of each duplex were measured and compared with the unmodified counterpart. The raw and relative thermodynamic measurements were then compared which indicated that differences occur that are both adduct and sequence dependent. These differences were next compared with the in vitro DNA polymerase incorporation data and were found to be strikingly correlated. Most significantly, for all four B[a]P isomers a mismatch of an A across from the adduct resulted in the least amount of relative destabilization, while the Watson-Crick complement C showed the most; in vitro studies showed that A is the preferred base incorporated across from each isomer, while C was incorporated least often. This observed correlation suggests that one factor contributing to misincorporation at an adduct site is the thermodynamic stability of the incorporated base. Structurally, the effect of sequence context and mismatched complementary strands were also compared, suggesting that all adducts tend to intercalate within the helix when they are complemented with a mismatched complementary strand. In addition, the level of this intercalation seems to be both sequence and stereoisomer dependent.