Stereochemical, structural, and thermodynamic origins of stability differences between stereoisomeric benzo[a]pyrene diol epoxide deoxyadenosine adducts in a DNA mutational hot spot sequence

Benzo[a]pyrene (BP), a prototype polycyclic aromatic hydrocarbon (PAH), can be metabolically activated to the enantiomeric benzo[a]pyrene diol epoxide s (BPDEs), (+)-(7R,8S,9S, 10R)-7,8-dihydroxy-9, 10-epoxy-7,8,9,10-tetrahydr obenzo[a]pyrene and the (-)-(7S,8R,9R,10S) enantiomer. These can react with adenine residues in DNA, to produce the stereoisomeric 10S (+)- and 10R (- )-trans-anti-[BP]-N-6-dA adducts. High-resolution NMR solution studies indi cate that in DNA duplexes the 10R (-) adduct is intercalated on the 5'-side of the modified adenine, while the 10S (+) adduct is disordered, exhibits multiple adduct conformations, and is positioned on the 3'-side of the modi fied adenine. Duplexes containing the 10S (+) adduct positioned at A*: with in codon 61 of the human N-ras sequence CAY:A are thermodynamically less st able and more easily excised by human DNA repair enzymes than those contain ing the 10R (-) adduct. However, the molecular origins of these differences are not understood and represent a fascinating opportunity for elucidating structure-function relationships. We have carried out a computational inve stigation to uncover the structural and thermodynamic origins of these effe cts in the 11-mer duplex sequence d(CGGACA*AGAAG). d(CTTCTTGTCCG) by perfor ming a 2-ns molecular dynamics simulation using NMR solution structures as the basis for the starting models. Then, we applied the MM-PBSA (molecular mechanics Poisson-Boltzmann surface area) method to compute free energy dif ferences between the stereoisomeric adducts. The 10R (-) isomer is more sta ble by similar to 13 kcal/mol, of which similar to 10 kcal/mol is enthalpic , which agrees quite well with their observed differences in thermodynamic stability. The lower stability of the 10S (+) adduct is due to diminished s tacking by the BP moiety in the intercalation pocket, more helix unwinding, and a diminished quality of Watson-Crick base pairing. The latter stems fr om conformational heterogeneity involving a syn-anti equilibrium of the gly cosidic bond in the modified adenine residue. The lower stability and confo rmational heterogeneity of the 10S (+) adduct may play a role in its enhanc ed susceptibility to nucleotide excision repair.