Mutagenic hot spots & 3-D structure of damaged DNA

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During the last project period, we used a single strand shuttle vector system to establish the mutagenic potential and specificity of selected DNA adducts. In addition, we developed a new procedure for the analysis of tamoxifen-DNA adducts and used it to demonstrate the genotoxicity of procedure for the analysis of tamoxifen-DNA adducts and used it to demonstrate the genotoxicity for this drug for the human endometrium. We reported that the adenine DNA glycosylase MutY forms a long-lived intermediate with DNA, thereby preventing double strand breaks during DNA repair. We cloned the mouse and human cognate genes for Ogg1 and showed that the gene product is a functional homolog of Fpg protein in bacteria. Future studies are designed to determine the mutagenic potential of selected environmental mutagens including bisphenol-A. We will use a new double strand vector that reveals the relative contribution of translesion synthesis, excision repair, and recombination events (damage tolerance) in the cellular processing of DNA damage in mammalian cells. We will also study mutation hot spots and related sequence context effects with respect to the reactivity proximate carcinogens with DNA. These experiments lay ground for future studies in molecular epidemiology in which adduct levels in humans are related to mutations observed in the P-53 gene. Using x-ray crystallographic techniques, we proposed to establish three dimensional structures of DNA containing defined lesions as a duplex oligonucleotide and as a ternary complex with rat liver DNA polymerase beta. These structural studies address our long-range goal of relating molecular structure and biological function.