Enzymology Of Eukaryotic Mismatch Repair

真核错配修复的酶学

基本信息

批准号：
7336327
负责人：
PAUL LAWRENCE MODRICH
金额：
$ 41.1万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1991
资助国家：
美国
起止时间：
1991-01-01 至 2010-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7336327
关键词：
Accounting Address Adenine Nucleotides Biochemical Biochemical Genetics Cells Cisplatin Clinical Colon Carcinoma Complex DNA DNA Binding DNA Damage DNA Repair DNA lesion DNA-Binding Proteins DNA-Directed DNA Polymerase Defect Development Ensure Enzymatic Biochemistry Excision Exodeoxyribonuclease I Genes Genetic Genetic Recombination Goals HMGB1 gene Human In Vitro Inherited Laboratories Lesion MLH1 gene MSH2 gene Mammalian Cell Mammals Mediating Mismatch Repair Molecular Molecular Conformation Mutation Nature Nuclear Extract PCNA gene Pathway interactions Pharmaceutical Preparations Phosphotransferases Play Preparation Process Proteins Reaction Replication Error Resistance Roentgen Rays Role SS DNA BP Signal Transduction Solutions Structure System Thioguanine Time Tissues Yeasts antitumor drug base cancer therapy cytotoxic interest novel reconstitution research study response stoichiometry tumor yeast two hybrid system

项目摘要

Mismatch repair is a major contributor to genetic stability. This pathway is of clinical interest because mutations in mismatch repair genes are the cause of one of the most prevalent forms of hereditary colon cancer, and have been implicated in the development of a subset (¿ 15%) of sporadic tumors that can occur in a variety of tissues. Mismatch repair defects also have implications for cancer treatment because inactivation of the pathway renders cells resistant to the cytotoxic effects of certain anti-tumor drugs. The primary goals of this project are to elucidate the molecular nature of this DNA repair system as it functions in replication error correction and in the cellular response to DNA damage. Our aims are four-fold: (1) Nine activities (MutSa, MutS(3, MutLa, Exol, RFC, PCNA, RPA, HMGB1, and DNA polymerase 6) have been implicated in eukaryotic mismatch repair, but it is clear that additional, and as yet unknown, components play important regulatory roles in the reaction. We intend to isolate these activities and establish their functions in the reaction. (2) We have shown that subsets of the known human mismatch repair activities catalyze several partial reactions that account at least in part for the complexity of the overall reaction. We plan further characterization of the corresponding multi-protein¿DNA assemblies with respect to molecular activities, stoichiometry, and conformation, as well as modulatory effects of adenine nucleotides on these parameters. (3) The structural basis of mismatch/DNA lesion recognition and processing by the human mismatch repair system will be addressed in collaborative X-ray crystallographic studies with Lorena Beese's laboratory. (4) MutSa and MutLa defects render mammalian cells resistant to the cytotoxic effects of SN1 DNA methylators, 6-thioguanine, and cisplatin. In an attempt to clarify the function of the human mismatch repair system in the DNA damage response, we will pursue in vitro experiments with the hope of elucidating the mechanism of MutSa- and MutLa-dependent activation of the ATR damage-signaling kinase in response to O6-methylguanine lesions.

不匹配的维修是遗传稳定性的主要因素。不匹配原因的突变是一些静脂结肠之一的原因癌症，并与可能发生的零星肿瘤的子集（15％）有关在多种组织中。途径的灭活使细胞对某些抗肿瘤药物的毒性作用产生该项目的主要目标是阐明该DNA CAIR系统在功能中的分子性质复制误差校正和对DNA损伤的细胞响应。活动（MUTSA，MUTS（3，MUTLA，EXOL，RFC，PCNA，RPA，HMGB1和DNA聚合酶6）与真核不匹配的克莱帕尔有关反应中的重要调节作用。已经表明，已将已知人类不匹配的子集撞倒催化我们计划的总体反应至少部分解释了一部分相关性多蛋白的进一步表征DNA组装相对于分子活动，化学计量和混乱，以及腺嘌呤核苷酸对这些的模块化效应参数。（3）人类的结构基础与Lorena Beese's的不匹配合作与Lorena Beese的不匹配合作的不匹配合作实验室。 DNA甲基，6-硫代氨酸和顺铂。 DNA损伤反应中的维修系统，我们将进行体外实验，希望阐明 ATR损伤信号激酶的MUTSA和MUTLA-MUTLA-EXTEPENTEPENT激活的机制响应至O6-甲基鸟氨酸病变。