CRYSTAL STRUCTURE OF DNA REPLICATION PROTEINS

DNA 复制蛋白的晶体结构

• 批准号: 6544644
• 负责人: JOHN KURIYAN
• 金额: $ 18.61万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-01-01 至 2002-12-09
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6544644
• 关键词: Archaea; DNA directed DNA polymerase; DNA replication; Escherichia coli; X ray crystallography; adenosinetriphosphatase; bacterial proteins; bacteriophage T4; cell cycle proteins; crystallization; enzyme mechanism; exonuclease; nucleic acid structure; oncoprotein p21; proliferating cell nuclear antigen; protein purification; protein structure; replicase; virus protein

DESCRIPTION (Adapted from abstract): X-ray crystallography will be used to study the structures of replicases (polymerase, sliding clamp protein, and complex that loads clamp onto DNA). Completed work has shown that the processivity factors from bacteria (beta subunit), eukaryotes (PCNA), and bacteriophage T4 (gene 45 protein) are ring-shaped proteins that share a commo architecture. These provide a sliding platform to which the polymerase machinery is tethered. Each of these replicases contains a multi-subunit clamp-loader. An active clamp-loader complex from E. coli (gamma, delta, and delta prime subunits) has been crystallized. An alternative strategy is to use proteins from thermophilic organisms. Several subunits of the replicase of the eubacterium Thermus thermophilis (T. th.) have been cloned. The DNA-dependent ATPase subunit of the clamp-loader of T. th. (gamma subunit) has been crystallized. The structure of the DNA polymerase/exonuclease from an extreme archaebacterial thermophile Desulfurucoccus tok, which is homologous to the PCNA-dependent mammalian DNA polymerases delta and epsilon, will be determined from crystals that diffract to 2.4 A. Although DNA replicases from various organisms share a common mechanistic framework, the protein components are quite divergent in sequence. The structural information resulting from this work may set the stage for structure-based design of replicase inhibitors with potential for high specificity.

描述（改编自摘要）：X 射线晶体学将用于 研究复制酶的结构（聚合酶、滑夹蛋白和 将夹子加载到 DNA 上的复合物）。 已完成的工作表明 来自细菌（β 亚基）、真核生物 (PCNA) 和 噬菌体 T4（基因 45 蛋白）是环状蛋白，具有相同的结构： 通用架构。 这些提供了一个滑动平台， 聚合酶机器被束缚。 这些副本中的每一个都包含一个 多子单元夹具装载机。 来自大肠杆菌的主动夹钳复合物 （gamma、delta 和 delta prime 亚基）已结晶。 一个 替代策略是使用来自嗜热生物的蛋白质。 嗜热栖热菌复制酶的几个亚基 （T.th.）已被克隆。 DNA 依赖性 ATP 酶亚基 T.th 的夹具装载机 （γ亚基）已结晶。 这 极端古细菌 DNA 聚合酶/核酸外切酶的结构 嗜热脱硫球菌 tok，与 PCNA 依赖型同源 哺乳动物 DNA 聚合酶 delta 和 epsilon，将由 衍射至 2.4 A 的晶体。尽管 DNA 复制品来自不同的 生物体具有共同的机制框架，蛋白质成分是 顺序上相差很大。 由此产生的结构信息 这项工作可能为基于结构的复制酶抑制剂设计奠定基础 具有高特异性的潜力。