FUNCTIONAL AND MECHANISTIC STUDIES OF DNA TOPOISOMERASES

DNA 拓扑异构酶的功能和机制研究

• 批准号: 2174990
• 负责人: LEROY F LIU
• 金额: $ 28.1万
• 依托单位: UNIV OF MED/DENT NJ-R W JOHNSON MED SCH
• 依托单位国家: 美国
• 财政年份: 1980
• 资助国家: 美国
• 起止时间: 1980-04-01 至 1997-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2174990
• 关键词: DNA; DNA repair; DNA replication; DNA replication origin; DNA topoisomerases; Drosophilidae; Escherichia coli; Saccharomyces cerevisiae; Xenopus; alternatives to animals in research; bacterial genetics; cell growth regulation; conformation; enzyme mechanism; enzyme structure; fungal genetics; gene expression; genetic promoter element; genetic recombination; laboratory mouse; laboratory rabbit; molecular cloning; molecular genetics; nucleic acid sequence; nucleic acid structure; simian virus 40; tissue /cell culture; transcription factor

The long-term objective of the proposed research is to understand the molecular mechanisms involved in regulation of DNA conformation. Regulation of DNA conformation is likely to be an integral part of genetic control mechanisms and its understanding may have broad implications in many areas of health-related genetic research. In the past, DNA topoisomerases have been identified as the major enzymes involved in regulating DNA conformation; More recently, accumulating evidence has indicated that significant local DNA conformational changes can be brought about by DNA helix-tracking processes such as transcription elongation and the action of certain DNA helix-tracking proteins including SV40 T antigen, E. coli UvrAB and some partially characterized DNA helix-tracking proteins from Xenopus laevis and Saccharomyces cerevisiae. In the present proposal, we will focus our studies on the regulation of local DNA conformation and its potential role in several genetic processes. A major part of the proposed research is to test whether the variable rates of movement of RNA polymerases along the DNA template can generate a high degree of local supercoiling which influences template functions in initiation of DNA replication, recombination and repair. We will also test whether anchored translocation is another means to regulate local DNA conformation. The possibility that E. coli UvrAB may alter local DNA conformation by a mechanism involving anchored translocation will be investigated. Finally, using an assay specific for DNA helix-tracking proteins, we will purify the DNA helix-tracking protein(s) from Saccharomyces cerevisiae and investigate its mechanism of translocation on duplex DNA.

拟议研究的长期目标是了解 参与 DNA 构象调节的分子机制。 DNA构象的调控可能是DNA构象的一个组成部分 遗传控制机制及其理解可能具有广泛的意义 对健康相关基因研究的许多领域的影响。 在 过去，DNA拓扑异构酶已被确定为主要酶 参与调节DNA构象；最近又在积累 有证据表明，显着的局部 DNA 构象变化 可以通过 DNA 螺旋追踪过程来实现，例如 转录延伸和某些 DNA 螺旋追踪的作用 蛋白质包括 SV40 T 抗原、大肠杆菌 UvrAB 和一些部分 鉴定了来自非洲爪蟾的 DNA 螺旋追踪蛋白， 酿酒酵母。 在本提案中，我们将重点关注 局部DNA构象调控及其潜力研究 在几个遗传过程中发挥作用。 拟议研究的主要部分 是为了测试RNA聚合酶的运动速率是否可变 沿着DNA模板可以产生高度的局部超螺旋 它影响 DNA 复制起始中的模板功能， 重组和修复。 我们还将测试是否锚定 易位是调节局部 DNA 构象的另一种方法。 这 大肠杆菌 UvrAB 可能通过以下方式改变局部 DNA 构象： 将研究涉及锚定易位的机制。 最后，使用特定于 DNA 螺旋追踪蛋白的检测方法，我们将 从酿酒酵母中纯化 DNA 螺旋追踪蛋白 并研究其在双链DNA上的易位机制。