This proposal involves (1) studies of the interactions of aflatoxin B1 with DNA and oligodeoxynucleotides, (2) development of facile methodology for site-specific introduction of deuterium into oligodeoxynucleotides, and (3) investigations of interactions of arylamines with DNA. Major goals of the proposed research are: 1. To establish, primarily through the use of NMR spectroscopy, the conformation and spatial orientation of chemically-induced covalent aflatoxin B1-oligodeoxynucleotide adducts, in particular the N7-guanyl adduct, and a primary degradation product, the formamidopyrimidine (FAPY) derivative. Covalent interactions at both week and strong binding sites will be investigated. Improved methodology for site-specific alkylation and purification of these adducts will be developed. 2. Investigations will be made of pre-covalent equilibrium binding interactions between aflatoxins and DNA, specifically as related to sequence and conformation of the DNA. Ultimately, specific non- covalent complexes will be compared with corresponding covalent adducts. 3. Investigations of the conformation and spatial orientation of photo-induced adducts. The aflatoxin photoadducts have received little attention but may provide a tool with which the conformations of non-covalently bound aflatoxins can be established. 4. Synthetic routes for site-specific incorporation of deuterium into various sites both in the heterocyclic bases and deoxyribose moieties of oligodeoxynucleotides will be developed. The primary focus of work in this area will be in the study of aflatoxin B1- DNA interactions by NMR. A route for complete deuteriation of the deoxyribose residue will be developed. 5. Investigations will be extended to other important environmental mutagens and carcinogens, in particular the electrophilic nitrogen species derived from aromatic amines.
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