Biochemical Analysis of Type II DNA Topoisomerases

II 型 DNA 拓扑异构酶的生化分析

• 批准号: 6852702
• 负责人: JAMES M BERGER
• 金额: $ 29.69万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-05-01 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6852702
• 关键词: DNA; DNA gyrase; X ray crystallography; adenosine triphosphate; antineoplastics; conformation; drug interactions; enzyme mechanism; enzyme structure; enzyme substrate complex; molecular cloning

DESCRIPTION (provided by applicant): The long-term objective of this proposal is to structurally define type II topoisomerase function and drug inhibition mechanisms. Type II topoisomerases are a ubiquitous class of proteins that use ATP to actively transport one DNA duplex through another. This reaction maintains appropriate levels of DNA supercoiling and resolves lethal chromosome tangles in the cell. Eukaryotic type II topoisomerases are also exploited by certain inhibitors that are frontline clinical therapies for cancer. The focus of this proposal is on eukaryotic topoisomerase II and archaeal topoisomerase VI, two representative members of type IIA and liB topoisomerases, respectively. The specific aims of the research are as follows: 1. To understand how type II topoisomerases physically switch between conformational states. 2. To define how type II topoisomerases interact with and utilize reaction substrates. 3. To determine how certain anticancer agents bind to and inhibit eukaryotic topoisomerase II Although a general framework exists for understanding type II topoisomerase function, significant gaps remain. A detailed structural description of type II topoisomerase mechanism coupled with biochemical validation, will illuminate critical aspects of these enzymes' function. The studies outlined here will define the physical events by which type II topoisomerases catalyze the ATP-dependent passage of one DNA segment through another and by which anticancer inhibitors subvert enzyme function. Data resulting from such efforts broadly impact a number of important scientific research fronts, from understanding how a molecular machine maintains chromosome topology to dissecting the molecular basis of various chemotherapeutic treatments.

描述（由申请人提供）： 该提案的长期目标是从结构上定义 II 型拓扑异构酶功能和药物抑制机制。 II 型拓扑异构酶是一类普遍存在的蛋白质，它们使用 ATP 主动将一个 DNA 双链体转运到另一个 DNA 双链体。该反应维持适当水平的 DNA 超螺旋并解决细胞中致命的染色体缠结。真核 II 型拓扑异构酶也被某些抑制剂所利用，这些抑制剂是癌症的一线临床疗法。该提案的重点是真核拓扑异构酶 II 和古菌拓扑异构酶 VI，分别是 IIA 型和 liB 型拓扑异构酶的两个代表性成员。 研究的具体目的如下： 1. 了解 II 型拓扑异构酶如何在构象状态之间进行物理切换。 2. 定义 II 型拓扑异构酶如何与反应底物相互作用并利用反应底物。 3. 确定某些抗癌药物如何结合并抑制真核拓扑异构酶 II 尽管存在理解 II 型拓扑异构酶功能的通用框架，但仍然存在重大差距。 II 型拓扑异构酶机制的详细结构描述与生化验证相结合，将阐明这些酶功能的关键方面。这里概述的研究将定义 II 型拓扑异构酶催化一个 DNA 片段通过另一个 DNA 片段的 ATP 依赖性通道以及抗癌抑制剂破坏酶功能的物理事件。这些努力产生的数据广泛影响了许多重要的科学研究前沿，从理解分子机器如何维持染色体拓扑到剖析各种化疗的分子基础。