This proposal addresses structure-dynamics-function relationships in DNA helices containing carcinogenic and mutagenic lesions at specific positions in oligonucleotides of defined sequence. We propose to combine two dimensional homonuclear and heteronuclear NMR methods with structure reconstruction algorithms to define features of the solution conformation of these DNA helices at and adjacent to the damage sites. The hydrogen exchange characteristics of the imino protons will be evaluated at infinite buffer catalyst concentration to estimate local base pair opening lifetimes at the individual base pair level. Specific projects include (1) characterization of oligonucleotide duplexes containing 04-and 02-alkylthymidine carcinogenic lesions ranging from methyl to isopropyl substituents. The base pairing potential of these alkylated pyrimidines with A and G will be probed to elucidate the origin of the observed transition and transversion errors associated with this lesion. (2) We propose to characterize structural and dynamics details at stable mutagenic abasic site lesions incorporated into oligonucleotide duplexes. Both furanose-intact and -open abasic sites will be investigated with A and T on partner strands opposite the lesion. Specific intermolecular interactions between tyrosine/tryptophan containing peptides and the abasic site will be characterized in detail. (3) The pairing properties of the mutagenic 2-aminopurine and 2,6-diaminopurine lesions with T and C will be monitored by proton and nitrogen-15 NMR parameters. We shall attempt to differentiate between Wobble base pairing and protonated base pair formation for the 2-aminopurine-cytosine interaction. (4) These mutagenic and carcinogenic lesions will be incorporated adjacent to consensus promotor regions involved in transcriptional regulation and adjacent to replication forks. The functional properties of regulatory sites as modulated by these lesions will be characterized by the proposed structural and hydrogen exchange experiments.