Structure of the 1,N2-etheno-2'-deoxyguanosine adduct in duplex DNA at pH 8.6.
The structure of the 1,N(2)-etheno-2'-deoxyguanosine (1,N(2)-epsilondG) adduct, arising from the reaction of vinyl chloride with dG, was determined in the oligonucleotide duplex 5'-d(CGCATXGAATCC)-3'.5'-d(GGATTCCATGCG)-3' (X=1,N(2)-epsilondG) at pH 8.6 using high resolution NMR spectroscopy. The exocyclic lesion prevented Watson-Crick base-pairing capability at the adduct site and resulted in an approximately 17 degrees C decrease in Tm of the oligodeoxynucleotide duplex. At neutral pH, conformational exchange resulted in spectral line broadening near the adducted site, and it was not possible to determine the structure. However, at pH 8.6, it was possible to obtain well-resolved (1)H NMR spectra. This enabled a total of 385 NOE-based distance restraints to be obtained, consisting of 245 intra- and 140 inter-nucleotide distances. The (31)P NMR spectra exhibited two downfield-shifted resonances, suggesting a localized perturbation of the DNA backbone. The two downfield (31)P resonances were assigned to G(7) and C(19). The solution structure was refined by molecular dynamics calculations restrained by NMR-derived distance and dihedral angle restraints, using a simulated annealing protocol. The generalized Born approximation was used to simulate solvent. The emergent structures indicated that the 1,N(2)-epsilondG-induced structural perturbation was localized at the X(6).C(19) base pair, and its 5'-neighbor T(5).A(20). Both 1,N(2)-epsilondG and the complementary dC adopted the anti conformation about the glycosyl bonds. The 1,N (2)-epsilondG adduct was inserted into the duplex but was shifted towards the minor groove as compared to dG in a normal Watson-Crick C.G base pair. The complementary cytosine was displaced toward the major groove. The 5'-neighbor T(5).A(20) base pair was destabilized with respect to Watson-Crick base pairing. The refined structure predicted a bend in the helical axis associated with the adduct site.
Shanmugam
Ganesh
G
Department of Chemistry, Vanderbilt Institute of Chemical Biology, Center in Molecular Toxicology, and the Vanderbilt-Ingram Cancer Center, Vanderbilt University, Nashville, Tennessee 37235, USA.
Goodenough
Angela K
AK
Kozekov
Ivan D
ID
Guengerich
F Peter
FP
Rizzo
Carmelo J
CJ
Stone
Michael P
MP
D004247:DNA / Q000737:chemistry*; D018736:DNA Adducts / Q000737:chemistry*; D004260:DNA Repair; D003839:Deoxyadenosines / Q000737:chemistry*; D003849:Deoxyguanosine / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D009690:Nucleic Acid Conformation; D011522:Protons
D018736:DNA Adducts; D003839:Deoxyadenosines; D011522:Protons; C032221:1,N(6)-ethenodeoxyadenosine; D004247:DNA; D003849:Deoxyguanosine