Structural characterization of a (+)-trans-anti-benzo[a]pyrene-DNA adduct using NMR, restrained energy minimization, and molecular dynamics.
The (+)-trans-anti-benzo[a]pyrene adduct formed at the N2 amino group of guanine is the major adduct found after metabolic activation of the ubiquitous carcinogen benzo[a]pyrene. The carcinogenic and mutagenic properties of the (+)-trans-anti-BP adduct, as well as related adducts, have been extensively studied. A DNA duplex containing a (+)-trans-anti-benzo[a]pyrene adduct covalently attached to the G8 nucleotide in the sequence d(CCTATGT[BP-G]CAC).d(GTGCACATAGG) was synthesized and the structure characterized by one- and two-dimensional NMR spectroscopy, in conjunction with energy minimization and molecular dynamics. This BP-11-mer duplex exhibits NOESY cross-peaks between benzo[a]pyrene protons and BP-G8, C9, A16, and C17 nucleotide protons that clearly delineate the location of the BP moiety in the minor groove of a B-type duplex with the pyrene ring oriented toward the 5' end of the modified strand. Large upfield shifts of A16 and C17 sugar resonances in the partner strand show that the pyrene moiety is situated near these sugars. Analysis of the spectra was complicated by the presence of chemical exchange line broadening of protons located near the (...T[BP-G]C...).(...GCA...) adduct site which shows the presence of a minor conformation for this BP-modified duplex in which TA is the 5' neighboring base pair. Distance restraints determined from NOESY spectra recorded at 20 degrees C were used in restrained and unrestrained energy minimization and molecular dynamics simulations to obtain a structure characteristic of the predominant conformation of the BP-11-mer duplex. The important structural features of the BP-11-mer are similar to those reported by Cosman et al. [(1992) Proc. Natl. Acad. Sci. U.S.A. 89, 1914-1918] for a (+)-trans-anti-BP adduct at a (...C[BP-G]C...).(...GCG...) sequence in which CG is the 5' neighboring base pair. No evidence of a conformational equilibrium was reported in this duplex, from which we conclude that the presence of a 5' TA base pair plays a role in the conformational equilibrium. Watson-Crick base pairing is retained in the predominant conformer of the (+)-trans-anti-BP modified duplex, which provides a visualization of a structure that could allow faithful replication. The exchange rate could not be slowed sufficiently to allow individual distance parameters to be obtained for the minor conformer.(ABSTRACT TRUNCATED AT 400 WORDS)
Fountain
M A
MA
Department of Chemistry, University of Rochester, New York 14627.
Krugh
T R
TR
D001483:Base Sequence; D001564:Benzo(a)pyrene / Q000737:chemistry*; D018736:DNA Adducts / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D011522:Protons; D013237:Stereoisomerism; D013816:Thermodynamics
D018736:DNA Adducts; D009838:Oligodeoxyribonucleotides; D011522:Protons; C056612:benzo(a)pyrene-DNA adduct; D001564:Benzo(a)pyrene