SPECIFICITY OF DNA REPAIR FOR OXIDIZED ABASIC SITES

氧化碱基位点 DNA 修复的特异性

• 批准号: 2700704
• 负责人: Bruce F. Demple
• 金额: $ 28.12万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-07-25 至 2000-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2700704
• 关键词: DNA damage; DNA repair; Escherichia coli; bacterial DNA; bacterial proteins; bleomycin; electrophoresis; endonuclease; enzyme activity; esters; exonuclease; gamma radiation; glycolates; high performance liquid chromatography; hydrogen peroxide; lactones; mutant; oxidation; oxidizing agents; sedimentation; yeasts

DESCRIPTION: Free radical damage to DNA, which has been associated with carcinogenesis, the normal aging process, and neurodegeneration, produces a variety of lesions. Many of these damages are various abasic lesions (sites of base loss). These include 1 -oxidized, 4 -oxidized or fragmented deoxyribose residues, known products of agents such as H2O2, ionizing radiation and bleomycin. Such damages have cytotoxic and mutagenic potential that might underlie the age and disease effects cited above, but these oxidized deoxyribose lesions have been difficult to measure specifically, sensitively and quantitatively by existing methods. This project proposes to develop new methods for such analysis, with an emphasis on the use of DNA repair enzymes to liberate specific damages for analysis by advanced techniques of mass spectrometry and capillary electrophoresis. These methods will be used to explore the formation and repair of individual oxidized abasic sites in vivo. Abasic (AP) endonucleases have been implicated in the repair of these damages, but this specificity has not been demonstrated in vivo. Key proposed studies include analyzing chromosomal DNA to determine the specific role of E. coli exonuclease III in repairing H2O2-induced DNA damage, and of E. coli endonuclease IV and yeast Apnl in the repair of oxidized abasic sites. The ability of human Ape endonuclease to correct specific repair defects in mutant strains of E. coli and yeast will be assessed by the novel methods, as will mutated forms of endonuclease IV and Apnl. Not only will this work be important for future work toward understanding the role of free-radical damage in aging, cancer, and neurodegeneration, but it will also provide quantum improvements in the methodology for molecular analysis of an important class of oxidative DNA damage.

描述：与DNA的自由基损害，与DNA有关 致癌作用，正常的衰老过程和神经退行性产生A 各种病变。 这些损害中有许多是各种卑鄙的病变（现场 基本损失）。 其中包括1个氧化，4个氧化或碎片 脱氧核糖残基，诸如H2O2等剂的已知产品，电离 辐射和博来霉素。 这样的损害具有细胞毒性和诱变 可能是上述年龄和疾病影响的潜力，但 这些氧化的脱氧核糖病变很难测量 具体而言，通过现有方法敏感和定量。 这 项目建议开发新的方法进行此类分析，重点 关于使用DNA修复酶解放特定的损害进行分析 通过质谱和毛细管电泳的晚期技术。 这些方法将用于探索个体的形成和修复 体内氧化的无骨位点。 abasic（ap）核酸内切酶已经 与这些损害的维修有关，但这种特异性尚未 在体内展示。 拟议的关键研究包括分析染色体 DNA确定大肠杆菌外切核酸酶III在修复中的特定作用 H2O2诱导的DNA损伤，以及大肠杆菌核酸内切酶IV和酵母APNL 修复氧化的无骨位点。 人猿核酸内切酶的能力 纠正大肠杆菌和酵母突变菌株中的特定修复缺陷 将通过新方法进行评估，而突变的核酸内切酶也会进行评估 IV和APNL。 这项工作不仅对将来的工作很重要 了解自由基损害在衰老，癌症和 神经变性，但它也将在 重要类氧化DNA的分子分析方法 损害。