REPAIR OF OXIDATIVELY DAMAGED GUANINES IN HUMAN

人体氧化损伤鸟嘌呤的修复

• 批准号: 6376818
• 负责人: A-Lien L Lu-Chang
• 金额: $ 22.26万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-14 至 2003-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6376818
• 关键词: DNA damage; DNA directed DNA polymerase; DNA repair; X ray; alkylating agents; aphidicolin; carcinogen testing; chemical carcinogen; chemical carcinogenesis; endonuclease; enzyme activity; guanine; human genetic material tag; human tissue; mutagen testing; mutagens; neoplasm /cancer genetics; nucleic acid sequence; oligonucleotides; oxidative stress; proliferating cell nuclear antigen; tissue /cell culture; ultraviolet radiation

Mismatch repair is an error avoidance pathway devoted to enhancing the fidelity of DNA replication and maintaining genetic stability. Mutations in human mismatch repair genes predispose individuals to cancer. Oxidative stress and metabolic processes which produce reactive oxygen species have been implicated as important causative agents of mutagenesis, carcinogenesis, aging, and a number of diseases. The human MutY homolog (hMYH) mismatch repair pathway for repairing A/G, A/C, and A/8-oxoG mismatches will be our major focus. The 8-oxoG lesion is a major stable product of DNA oxidative damage and has the most deleterious effects because it can mispair with adenine. Thus, A/8-oxoG mismatches are particularly important biological substrates for hMYH. hMYH, like the E. coli MutY protein, is an adenine DNA glycosylase. Because recombinant hMYH protein expressed in E. coli and native hMYH have different mismatch specificities, the structural and functional differences of these proteins will be further analyzed. Glycosylase and apurinic/apyrimidinic (AP) lyase activities on DNA containing A/G, A/C, A/8-oxoG, and other base analogs will be assayed. Our results indicate the hMYH, MutS homologs (hMSH2 and hMSH6), and MutL homologs (hMLH1 and hPMS2) are associated with the DNA replication complex, thus the interactions between hMYH and replicative as well as mismatch repair proteins will be investigated by co-immunoprecipretation and affinity chromatography. The coupling of hMYH repair with DNA replication may direct MYH repair to the misincorporated adenines on the daughter strands. The effect of proliferating cell nuclear antigen (PCNA, an accessory factor for DNA polymerases delta and epsilon) on hMYH activity will be determined. Human breast and lung cancer cells will be analyzed for hMYH expression and screened for mutations in the hMYH gene. The sensitivities of the MYH defective cells to oxidative agents and radiation will be analyzed. Through the study of the mechanism of DNA mismatch repair, our understanding of cancer, aging, and genetic diseases can be advanced.

不匹配维修是一种避免途径，致力于增强 DNA复制的保真度和维持遗传稳定性。 人不匹配修复基因的突变使个体倾向于 癌症。 产生反应性的氧化应激和代谢过程 氧物种已被认为是重要的原因。 诱变，癌变，衰老和许多疾病。 人类 Muty同源物（HMYH）不匹配修复途径用于修复A/G，A/C和 A/8-oxog不匹配将是我们的主要重点。 8-oxog病变是 DNA氧化损伤的主要稳定产物，并且具有最多的 有害影响是因为它会与腺嘌呤混乱。 因此，a/8-oxog 不匹配是HMYH特别重要的生物底物。 HMYH与大肠杆菌型蛋白一样，是腺嘌呤DNA糖基酶。 因为重组HMYH蛋白在大肠杆菌和天然HMYH中表达 具有不同的不匹配特异性，结构和功能 这些蛋白质的差异将进一步分析。 糖基酶和 含有A/G，A/C的DNA上的Apurinic/Apyrimidinic（AP）裂解酶活性 A/8-oxog和其他基本类似物将被分析。 我们的结果表明 HMYH，MUTS同源物（HMSH2和HMSH6）和MUTL同源物（HMLH1和 HPMS2）与DNA复制复合物有关，因此 HMYH与复制和不匹配维修之间的相互作用 蛋白质将通过共免疫沉淀和亲和力研究 色谱法。 HMYH修复与DNA复制的耦合可能 将MYH维修直接到女儿的衰老腺丝 链。 增殖细胞核抗原的影响（PCNA，A DNA聚合酶Delta和Epsilon的辅助因子）在HMYH活性上 将确定。 将分析人类乳腺癌和肺癌细胞 用于HMYH表达并筛选HMYH基因中的突变。这 MYH缺陷细胞对氧化剂的敏感性和 辐射将进行分析。 通过研究DNA机制 不匹配维修，我们对癌症，衰老和遗传的理解 疾病可以采取病情。