title: NMR study of stacking interactions and conformational adjustments in the dinucleotide-carcinogen adduct 2'-deoxycytidylyl-(3----5)-2'-deoxy-8-(N-fluoren-2-ylac etamido)guanosi ne.
abstract: The conformation and dynamics of the dinucleotide d-CpG modified at the C(8) position of the guanine ring by the carcinogen 2-(acetylamino)fluorene has been investigated by high-field 1H NMR spectroscopy. A two-state analysis of chemical shift data has enabled estimation of the extent of intramolecular stacking in aqueous solution as a function of temperature. The stacking, which is mostly fluorene-cytosine, is virtually complete in the low-temperature range. The 500-MHz 1H NMR spectrum consists of two subspectra near ambient temperatures due to a 14.3 +/- 0.3 kcal/mol barrier to internal rotation about the amide bond in the stacked form. A large barrier to internal rotation about the guanyl-nitrogen bond at C(8) has also been ascertained, but separate NMR subspectra were not detected due to the predominance of one of the torsional diastereomers (alpha' = 90 degrees) in the fully stacked state. Problems of self-association and chemical exchange were identified and overcome to enable analysis of the sugar-phosphate backbone conformation utilizing coupling constants. For the exocyclic C(4')-C(5') bond of the deoxyguanosine moiety, there is a high gauche+ (gamma = 60 degrees) conformer population, which is uncommon for a purine nucleotide with a syn orientation about the glycosyl bond. The gauche- conformation (gamma = 300 degrees), which is normally present in syn purine nucleotides in solution, was not detected. The exocyclic C(5')-O(5') torsion of the deoxy-guanosine moiety remains near the classical energy minimum (beta = 180 degrees) in the major stacked conformations. The sugar ring of the deoxycytidine moiety is predominantly in the C2'-endo conformation, while the deoxyguanosine ring is a mixture of conformations, one of which appears to be unusually puckered. The results support intercalation models of modified DNA and suggest a looped-out structure, with the modified guanine being the first base in the loop. Such structures could explain the relatively rapid rate of repair and the frame-shift mutations of this type of adduct.
authors.affiliation: Division of Biochemical Toxicology (HFT-110), Food and Drug Administration, Jefferson, Arkansas 72079.
authors.forename: F E
authors.initials: FE
authors.lastname: Evans
authors.forename: R A
authors.initials: RA
authors.lastname: Levine
mesh_terms: D015073:2-Acetylaminofluorene; D003843:Deoxycytidine Monophosphate / Q000031:analogs & derivatives / Q000138:chemical synthesis; D003845:Deoxycytosine Nucleotides / Q000138:chemical synthesis; D003849:Deoxyguanosine / Q000031:analogs & derivatives*; D015226:Dinucleoside Phosphates; D006151:Guanosine / Q000031:analogs & derivatives* / Q000138:chemical synthesis; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D009690:Nucleic Acid Conformation; D013816:Thermodynamics
chemical_list: D003845:Deoxycytosine Nucleotides; D015226:Dinucleoside Phosphates; D003843:Deoxycytidine Monophosphate; D006151:Guanosine; C037781:deoxycytidylyl-(3'-5')-deoxyguanosine; C056751:2'-deoxycytidylyl-(3--5)-2'-deoxy-8-(N-fluoren-2-ylacetamido)guanosine; D015073:2-Acetylaminofluorene; D003849:Deoxyguanosine