Mutagenic hot spots & 3-D structure of damaged DNA

诱变热点

• 批准号: 6301315
• 负责人: SHINYA SHIBUTANI
• 金额: $ 21.9万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-10 至 2001-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6301315
• 关键词: DNA damage; DNA directed DNA polymerase; DNA repair; X ray crystallography; adduct; chemical carcinogen; environmental toxicology; mutagens; nucleic acid sequence; nucleic acid structure; oligonucleotides; phenolic amine

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.

在上一个项目期间，我们使用单链穿梭载体系统来确定所选DNA加合物的致突变潜力和特异性。此外，我们开发了一种分析他莫昔芬-DNA加合物的新方法，并用于证明他莫昔芬-DNA加合物分析方法的遗传毒性，并用于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糖基化酶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Ogg 1的小鼠和人类同源基因，并表明该基因产物是细菌中Fpg蛋白的功能同源物。未来的研究旨在确定选定的环境诱变剂（包括双酚A）的致突变潜力。我们将使用一种新的双链载体，揭示了在哺乳动物细胞中DNA损伤的细胞加工中translesion合成，切除修复和重组事件（损伤耐受性）的相对贡献。我们还将研究突变热点和相关的序列背景效应与DNA的反应性接近致癌物。这些实验奠定了基础，在分子流行病学中，在人类中的加合物水平与P-53基因中观察到的突变的未来研究。使用X-射线晶体学技术，我们建议建立三维结构的DNA含有定义的病变作为一个双链体寡核苷酸和作为一个三元复合物与大鼠肝DNA聚合酶β。这些结构研究解决了我们将分子结构与生物功能联系起来的长期目标。