Solution Structure of the Minor Conformer of a DNA Duplex Containing a dG Mismatch Opposite a Benzo[a]pyrene Diol Epoxide/dA Adduct:  Glycosidic Rotation from Syn to Anti at the Modified Deoxyadenosine

Abstract

Polycyclic aromatic hydrocarbons (PAHs) are widespread environmental contaminants whose metabolism in mammals results in deleterious cell transformation. Covalent modification of DNA by diol epoxides metabolically formed from PAHs such a benzo[a]pyrene (BaP) provides a mechanism for the genotoxicity, mutagenicity, and carcinogenicity of PAHs. We had previously reported NMR evidence for a minor conformer of the duplex d(G1G2T3C4A5*C6G7A8G9).d(C10T11C12G13G14G15A16C17C18) containing a dG14 mismatch opposite a dA5* residue modified at the exocyclic amino group by trans addition to (+)-(7R,8S,9S,10R)-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a] pyrene [Yeh, H.J.C., Sayer, J.M., Liu, X., Altieri, A.S., Byrd, R.A., Lashman, M.K., Yagi, H., Schurer, E.J., Gorenstein, D.G., & Jerina, D.M. (1995) Biochemistry 34, 13570-13581]. In the present work, we describe the structure of this minor conformer (ca. 17% of the total conformer population). This represents the first structural determination of a minor conformer of a carcinogen-lesion DNA adduct. Two-dimensional NOESY, ROESY, TOCSY, and exchange-only spectra at 750 MHz allowed nearly complete sequential assignment of both conformers. In the minor conformer, the adducted base assumes an anti-glycosidic torsion angle whereas in the major conformer it assumes an unusual syn-glycosidic torsion angle. The aromatic hydrocarbon in the minor conformer is intercalated between dG13 and dG14, preserving the energetically favorable stacking interactions found in the major conformer. The major structural differences between the two conformers appear to be near the lesion site as evidenced by the large chemical shift differences between major and minor conformer protons near the lesion site; away from this site, the chemical shifts of the major and minor conformer protons are nearly identical. Because any of the conformations of benzo[a]pyrene diol epoxide-modified DNA may contribute to tumorigenic activity, structural determination of all conformations is essential for the elucidation of the mechanism of cell transformation initiated by covalent modification of DNA by PAHs.

Thermodynamic and structural factors in the removal of bulky DNA adducts by the nucleotide excision repair machinery

“…20,21 The 10R(Ϫ)-trans-anti adduct structure shown is essentially the same as that of the NMR solution structure of Zegar et al 22 determined in the same C[A*]A sequence context of codon 61 of the human N-ras gene.…”

“…NMR solution structures provided starting models for these simulations. 16,17,21,22 gives rise to multiple and poorly resolved NMR resonances, as well as line broadening, all of which preclude a determination of a high-resolution structure for this stereoisomeric adduct. 21,[23][24][25] However, when the T complementary to the 10S(ϩ)-trans-anti-B[a]P-N 6 -dA is replaced by a mismatched G two different conformations were discerned and characterized.…”

“…16,17,21,22 gives rise to multiple and poorly resolved NMR resonances, as well as line broadening, all of which preclude a determination of a high-resolution structure for this stereoisomeric adduct. 21,[23][24][25] However, when the T complementary to the 10S(ϩ)-trans-anti-B[a]P-N 6 -dA is replaced by a mismatched G two different conformations were discerned and characterized. Although the major conformer has the glycosidic torsion angle, , at the modified adenine in the syn conformation, and the minor conformer has in the anti conformation, 21,24 in both cases the aromatic B[a]P residue is oriented on the 3Ј-side of the lesion, as it is in the stereochemically related 1S(Ϫ)-trans-…”

“…5 Because of this absence of an experimental NMR solution structure for the 10S(ϩ)-trans-anti adduct with normal partner dT opposite the lesion, molecular dynamics simulations were employed to elucidate their structures. 19 The NMR solution structure of Schwartz et al 21 in a duplex with an A*:G mismatch was remodeled 19 to the human N-ras codon 61 sequence I, with fully complementary A*:T base pair at the lesion site (Figure 4). The anti conformer for the glycosidic torsion angle was employed as the starting model because the normal partner dT was investigated.…”

Highly Mutagenic Bypass Synthesis by T7 RNA Polymerase of Site-specific Benzo[a]pyrene Diol Epoxide-adducted Template DNA

“…It has been shown that the stability of a (ϩ)-anti-trans-BPDE-adducted adenine mismatched with dG is significantly higher than the stability of the adducted adenine paired with the correct dT (41,43). This suggests that the differences in stability between the incorrect and correct base pairs containing the different diastereomers may play a role in BPDEinduced mutagenesis.…”