RECOGNITION AND REPAIR OF UV DAMAGE TO HUMAN DNA

人类 DNA 紫外线损伤的识别和修复

• 批准号: 6377972
• 负责人: Yue Zou
• 金额: $ 24.9万
• 依托单位: EAST TENNESSEE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-07-01 至 2005-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6377972
• 关键词: DNA damage; DNA footprinting; DNA repair; adduct; benzopyrenediol epoxide; chemical carcinogenesis; chemical kinetics; deoxyribonuclease I; enzyme activity; fluorescence resonance energy transfer; fluorescence spectrometry; gel mobility shift assay; gene mutation; human genetic material tag; neoplasm /cancer genetics; radiation carcinogenesis; site directed mutagenesis; thermodynamics; ultraviolet radiation

Individuals who suffer from the genetic disease xeroderma pigmentosum (XP) lack nucleotide excision repair (NER) of DNA, and thus have much higher carcinogenic probability than the average population after exposure to UV irradiation. NER is a major cellular biological defense system to remove DNA damage due to the formation of bulky lesions induced by UV irradiation and environmental genotoxic chemicals and carcinogens. Although, it has been generally accepted that DNA damage recognition plays a central role in NER, the molecular and thermodynamic details of DNA damage processing remain largely unclear, and have not been systematically studied using biochemically rigorous approaches. In addition, the roles of the damage recognition proteins XPA, RPA, and XPC-HR23B have been controversial. The long-term objective of this study is to understand the molecular and biochemical details of UV-induced or related DNA damage recognition and repair by human NER proteins, and the potential effects of these relationships on damage-induced mutagenesis and carcinogenesis. To gain a systematic and more precise view of DNA damage recognition and repair by human NER, the following questions will be addressed: What is the hierarchy of damage recognition and how is damage dynamically processed in a stepwise recognition mechanism? What structural and chemical alterations in the DNA helix are identified by repair proteins at specific recognition steps? What is the molecular architecture of recognition intermediates? And what protein domains are important for protein-DNA and protein-protein contacts in the recognition. Specifically, this project aims to determine the thermodynamics and kinetics of the interaction of XPC-HR23B, XPA, and RPA with DNA substrates containing site-specific UV photolesions and benzo[a]pyrene diol epoxide (BPDE) DNA adducts using rigorous biochemical approaches: identify and analyze the important protein motifs involved in damage recognition: characterize the repair intermediates for damage recognition of UV-induced photolesions and BPDE-DNA adducts; and determine the mechanism in which the structural and chemical modifications of damage are recognized and repaired.

患有遗传性色素性干皮病的个人 (XP)缺乏DNA的核苷酸切除修复(NER)，因此有很多 暴露后致癌概率高于一般人群 紫外光照射。NER是一个主要的细胞生物防御系统，需要移除 紫外线照射引起的块状损伤引起的DNA损伤 环境中的遗传毒性化学品和致癌物。尽管，它一直是 人们普遍认为DNA损伤识别在NER中起着核心作用， DNA损伤处理的分子和热力学细节在很大程度上仍然存在 还不清楚，而且还没有用严格的生化方法系统地研究过 接近了。此外，损伤识别蛋白XPA、RPA、 和XPC-HR23B一直存在争议。这项研究的长期目标是 了解紫外线诱导或相关的分子和生化细节 人类NER蛋白对DNA损伤的识别和修复及其潜力 这些关系对损伤诱导的突变和 致癌。为了获得对DNA损伤的系统和更准确的看法 对于人类NER的识别和修复，将解决以下问题： 损害识别的层次是什么？如何动态地识别损害 在一种逐步识别机制中处理？什么结构和化学物质 DNA螺旋的改变是由特定的修复蛋白识别的 认可步骤？识别的分子结构是什么？ 中间人？以及哪些蛋白质结构域对蛋白质-DNA和 蛋白质-蛋白质接触中的识别。具体地说，该项目旨在 测定XPC-HR23B、XPA、XPC相互作用的热力学和动力学 和带有DNA底物的RPA，该DNA底物包含特定部位的紫外光刻和 苯并[a]芘二醇环氧化物(BPDE)DNA加合物的严格生化研究 方法：确定和分析参与的重要蛋白质基序 损坏识别：对损坏的修复中间体进行表征 紫外光诱导的光刻和BPDE-DNA加合物的识别；并确定 损伤的结构和化学修饰是 已被识别并修复。