Biochemical Mechanism of DNA Polymerase III Holoenzyme

DNA聚合酶III全酶的生化机制

• 批准号: 8706000
• 负责人: Michael O'Donnell
• 金额: $ 10万
• 依托单位: Joan and Sanford I. Weill Medical College of Cornell University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-08-01 至 1989-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=8706000&HistoricalAwards=false
• 关键词: Biochemical; Mechanism; DNA; Polymerase; III

DNA polymerase III holoenzyme (Pol1III holoenzyme), the major replicative polymerase of E. coli is isolated as a complex of at least eight polypeptides and can be reconstituted from purified preparations of subunits. Although individual functions of some of the subunits are still unknown, collectively they confer upon the pol1III holoenzyme special properties that distinguish it as a major replicative polymerase. It is extremely rapid in locating a primer, it hydrolyzes ATP to initiative extremely processive synthesis, has high catalytic efficiency and diffuses on duplex DNA to search for a primer terminus. In this proposal the specific aims are to study the diffusion properties of the enzyme on DNA, the mechanisms underlying rapid location of a primer terminus, the stoichiometry and spatial orientation of subunits within the enzyme and their interaction with a primer terminus. The ultimate aim of this program is a detailed molecular description of the events at the replication fork of a growing chromosome and a grasp of the basic principles that underlie them. Dr. O'Donnell has received excellent postdoctoral training with Kornberg and with Lehman, and is skilled in the techniques of biochemistry and molecular biology. He is well qualified to pursue the proposed research.

DNA聚合酶III全酶(Pol1III全酶)是大肠杆菌的主要复制聚合酶，由至少8个多肽组成的复合体，可从纯化的亚基制剂中重组。尽管一些亚基的个别功能尚不清楚，但总的来说，它们赋予了pol1III全酶特殊的性质，使其成为主要的复制聚合酶。它极快地找到了一条引物，它能将三磷酸腺苷水解酶催化合成，具有很高的催化效率，并能扩散到双链DNA上寻找一条引物的末端。在这一建议中，具体目的是研究酶在DNA上的扩散特性，快速定位引物末端的机制，酶中亚基的化学计量比和空间取向，以及它们与引物末端的相互作用。这一计划的最终目的是详细的分子描述正在生长的染色体复制分叉上的事件，并掌握它们背后的基本原理。O‘Donnell博士在Kornberg和Lehman接受过出色的博士后培训，精通生物化学和分子生物学技术。他完全有资格从事拟议中的研究。