Mutagenic hot spots & 3-D structure of damaged DNA

在最后一个项目期间，我们使用单链穿梭载体系统来建立所选DNA加合物的诱变潜力和特异性。此外，我们开发了一种新的他莫昔芬-DNA加合物分析方法，并用它来证明他莫昔芬-DNA加合物分析程序的遗传毒性，并用它来证明该药物对人子宫内膜的遗传毒性。我们报道了腺嘌呤DNA糖基酶MutY与DNA形成一个长寿命的中间体，从而防止DNA修复过程中的双链断裂。我们克隆了Ogg1的小鼠和人类同源基因，结果表明该基因产物是细菌中的FPG蛋白的功能同源物。未来的研究旨在确定包括双酚-A在内的选定环境诱变剂的诱变潜力。我们将使用一种新的双链载体来揭示跨损伤合成、切除修复和重组事件(损伤耐受)在哺乳动物细胞DNA损伤的细胞处理中的相对贡献。我们还将研究突变热点和相关的序列上下文效应，涉及与DNA具有反应性的最接近的致癌物。这些实验为未来的分子流行病学研究奠定了基础，在分子流行病学中，人类的加合物水平与观察到的P-53基因突变有关。利用X射线结晶学技术，我们建议建立含有明确损伤的DNA的三维结构，作为双链寡核苷酸和大鼠肝DNA聚合酶β的三元复合体。这些结构研究解决了我们将分子结构和生物功能联系起来的长期目标。

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.