Solution structure of the 2-amino-1-methyl-6-phenylimidazo [4,5-b]pyridine (PhIP)-C8-dG adduct opposite dC in a DNA duplex

K Brown,PhD, a E Guenthera, PhD,a BE Hingerty,PhD,b S Broyde, PhD,c K Turteltaub,PhD,a M Cosman,PhD,a

aBiology & Biotechnology Research Program, Lawrence Livermore National Laboratory, Livermore CA bLife Sciences Division, Oak Ridge National Laboratory, Oak Ridge, TN cBiology Dept, New York University, New York, NY

The heterocyclic amine PhIP is a mutagenic compound that is formed during the cooking of fish and meats and has been shown to pose a significant dietary cancer risk to humans. We have used a combined NMR-molecular mechanics computational approach to determine the solution structure of the major C8-dG adduct formed by reaction of N-acetoxy-PhIP with an 11mer duplex sequence. The NMR data show that the [PhIP]dG.dC 11mer duplex undergoes a slow conformational exchange in which the PhIP ligand either intercalates between the flanking dG.dC base pairs by displacing the modified dG residue into the major groove, or is located outside the helix and solvent exposed in a minimally perturbed B-DNA duplex. The amount of the external binding conformer present is temperature dependent and ranges from ~10% at 25 oC, 15% at 20 oC and 32% at 1 oC. The PhIP phenyl ring undergoes an additional fast rotation with respect to the imidazo-pyridine fused ring system. However, the phenyl ring is clearly forming stacking interactions with the deoxyribose sugar protons of the flanking dG residues on the complementary strand in the main base-displaced intercalation structure, as evidenced by the unusually shifted proton resonances of these sugar protons due to the phenyl ring current effects contributions. Both proton and carbon chemical shift data for the sugar resonances of the PhIP modified dG are consistent with a syn glycosidic torsion angle for this residue, which is directed toward the 3'end of the modified strand. The conformational equilibrium observed for the PhIP-C8-dG adduct may, in part, explain the variety of the types of mutations induced by this adduct.

KEY WORDS: heterocyclic amine, solution structure, computational biology, mutagen.

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Dietary Influences.

This presentation received an honorable mention in our poster contest and was recognized with the Symposium Presidents' Award for Scientific Excellence.