Function of the Six ATPases in the Yeast 19S Particle

酵母 19S 颗粒中六种 ATP 酶的功能

• 批准号: 7060314
• 负责人: Gary L. Kleiger
• 金额: $ 5.2万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-04-01 至 2007-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7060314
• 关键词: adenosine triphosphate; adenosinetriphosphatase; analog; chemical genetics; enzyme activity; enzyme inhibitors; enzyme substrate; fungal genetics; fungal proteins; gene expression; gene mutation; intracellular transport; membrane transport proteins; microorganism culture; postdoctoral investigator; proteasome; protein binding; protein degradation; protein folding; protein structure function; protein transport; site directed mutagenesis; ubiquitin; yeasts

DESCRIPTION (provided by applicant): The elucidation of the ubiquitin/proteasome system in eukaryotes has greatly contributed to our understanding of how cells work. However, many mechanistic features of how 26S proteasomes function remain unclear. One key question involves the function of the six AAA ATPases found in the 19S regulatory particle of the proteasome. Despite the importance of these proteins in proteasome function, very few studies have directly addressed the specific roles of ATPase activity in proteasome function. The goal of this proposal is to apply a novel 'chemical genetics' approach that will enable us to systematically deduce the molecular function of each ATPase in the 19S. Inhibitors are generated by chemically modifying purine derivatives with bulky substituents to complement a rationally designed enlargement of the ATP-binding sites of the target ATPases. While the 'analogue-sensitive' ATPases can bind to either normal ATP or the ATP analogue, wild-type ATPases are unaffected in the presence of ATP analogue, allowing specific inhibition of a given ATPase in both in vitro and in vivo assays. Initially we will characterize the roles of the yeast 19S ATPases Rpt1 and Rpt5 in proteasome function, with the goal of eventually characterizing all six ATPases.

描述（由申请人提供）：真核生物中泛素/蛋白酶体系统的阐明极大地促进了我们对细胞如何工作的理解。然而，26S 蛋白酶体如何发挥作用的许多机制特征仍不清楚。一个关键问题涉及蛋白酶体 19S 调节颗粒中发现的六种 AAA ATP 酶的功能。尽管这些蛋白质在蛋白酶体功能中很重要，但很少有研究直接探讨 ATP 酶活性在蛋白酶体功能中的具体作用。该提案的目标是应用一种新颖的“化学遗传学”方法，使我们能够系统地推断 19S 中每种 ATP 酶的分子功能。抑制剂是通过用大量取代基对嘌呤衍生物进行化学修饰来产生的，以补充合理设计的目标 ATP 酶 ATP 结合位点的扩大。虽然“类似物敏感”ATP 酶可以与正常 ATP 或 ATP 类似物结合，但野生型 ATP 酶在 ATP 类似物存在的情况下不受影响，从而可以在体外和体内测定中特异性抑制给定的 ATP 酶。首先，我们将表征酵母 19S ATP 酶 Rpt1 和 Rpt5 在蛋白酶体功能中的作用，目标是最终表征所有六种 ATP 酶。