REPAIR OF OXIDATIVELY DAMAGED GUANINES IN HUMAN

人体氧化损伤鸟嘌呤的修复

• 批准号: 6513267
• 负责人: A-Lien L Lu-Chang
• 金额: $ 22.93万
• 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/6513267
• 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.

错配修复是一种错误避免途径，致力于增强 DNA复制的保真度和维持遗传稳定性。 人类错配修复基因的突变使个体易患 癌 氧化应激和代谢过程产生反应性 氧类已经被认为是 诱变、致癌、衰老和许多疾病。 人类 MutY同源物（hMYH）错配修复途径，用于修复A/G、A/C和 A/8-oxoG错配将是我们的主要焦点。 8-oxoG损伤是 DNA氧化损伤的主要稳定产物， 因为它可以与腺嘌呤错配。 因此，A/8-oxoG 错配是hMYH特别重要的生物底物。 hMYH，就像E。大肠杆菌MutY蛋白是一种腺嘌呤DNA糖基化酶。 由于重组hMYH蛋白在E. coli和天然hMYH 具有不同的错配特异性， 将进一步分析这些蛋白质的差异。 糖基化酶和 无嘌呤/无嘧啶（AP）裂解酶对含有A/G、A/C、 将测定N8-oxoG和其他碱基类似物。 我们的研究结果表明 hMYH、MutS同源物（hMSH 2和hMSH 6）和MutL同源物（hMLH 1和hMLH 6） hPMS 2）与DNA复制复合物相关，因此 hMYH与复制以及错配修复之间的相互作用 蛋白质将通过共免疫识别和亲和性来研究 层析 hMYH修复与DNA复制的偶联可能 MYH直接修复了女儿的错误腺嘌呤 股。 增殖细胞核抗原（PCNA， DNA聚合酶δ和δ的辅助因子）对hMYH活性的影响 将被确定。 人类乳腺癌和肺癌细胞将被分析 用于hMYH表达并筛选hMYH基因中的突变。的 MYH缺陷细胞对氧化剂的敏感性， 将对辐射进行分析。 通过对DNA作用机制的研究， 错配修复，我们对癌症，衰老和遗传的理解 疾病可以发展。