CAREER: Protein Unfolding by Energy Dependent Proteases

职业：通过能量依赖性蛋白酶展开蛋白质

• 批准号: 9875857
• 负责人: Andreas Matouschek
• 金额: $ 50万
• 依托单位: Northwestern University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-02-01 至 2004-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9875857&HistoricalAwards=false
• 关键词: CAREER; Protein; Unfolding; Energy; Dependent

9875857The goal of this project is to determine the mechanism of unfolding of substrate proteins during degradation by ATP-dependent proteases. Most protein degradation in cells outside the vacuole or the lysosome is catalyzed by ATP-dependent proteases. Unfolding of the substrate protein is a critical early step in the degradation process. Degradation of short-lived proteins is essential for a wide range of cellular functions including cell cycle control, signal transduction and antigen presentation. Proteolysis of misfolded proteins is part of the cellular response to stress. Barnase, whose folding is well characterized in vitro, was chosen as an artificial substrate protein for the ATP-dependent proteases lon-protease, Clp-protease, mitochondrial AAA-protease and the proteasome. Barnase unfolding during degradation will be compared to spontaneous unfolding to determine whether protein unfolding is catalyzed by the proteases. Destabilized barnase mutants will be used as models for misfolded proteins and barnase mutants with C-terminal degradation tags will be used as models for short-lived regulatory proteins. If unfolding is catalyzed, the mechanism of catalysis will be determined. The educational aim of this award is to teach undergraduate students the prerequisites for a biochemical and biophysical understanding of cell biological processes. For this purpose a laboratory course on quantitative biophysical techniques will be developed.Lay summaryProteins are chains made from subunits called amino acids. To perform their biological function, proteins fold into a specific three-dimensional shape. However, there are essential processes in the cell for which proteins must unfold again, such as when damaged or regulatory proteins are degraded by proteases. Failure to degrade regulatory proteins and misfolded proteins leads to abnormal cellular function. This proposal investigates how proteases unfold proteins.Biology has reached a stage where it is possible to determine thedetailed mechanisms of processes in the cell. To prepare students for this new era in experimental biology, a laboratory course on quantitative methods in cell biology will be developed.

9875857本项目的目的是确定底物蛋白在ATP依赖性蛋白酶降解过程中的解折叠机制。 大多数蛋白质降解细胞外的液泡或溶酶体是由ATP依赖性蛋白酶催化。 底物蛋白的解折叠是降解过程中的关键早期步骤。 短寿命蛋白质的降解对于广泛的细胞功能包括细胞周期控制、信号转导和抗原呈递是必需的。 错误折叠蛋白质的蛋白水解是细胞对应激反应的一部分。 Barnase是一种在体外具有良好折叠特性的蛋白酶，被选择作为ATP依赖性蛋白酶离子蛋白酶、Clp蛋白酶、线粒体AAA蛋白酶和蛋白酶体的人工底物蛋白。将降解期间的芽孢杆菌RNA酶解折叠与自发解折叠进行比较，以确定蛋白质解折叠是否由蛋白酶催化。 失稳芽孢杆菌RNA酶突变体将用作错误折叠蛋白的模型，具有C-末端降解标签的芽孢杆菌RNA酶突变体将用作短寿命调节蛋白的模型。 如果解折叠被催化，则催化机制将被确定。 该奖项的教育目的是教本科生的先决条件，为细胞生物学过程的生物化学和生物物理的理解。 为此，将开设一门关于定量生物物理技术的实验课。蛋白质是由称为氨基酸的亚基组成的链。 为了发挥其生物学功能，蛋白质折叠成特定的三维形状。 然而，在细胞中存在蛋白质必须再次展开的基本过程，例如当受损或调节蛋白质被蛋白酶降解时。不能降解调节蛋白和错误折叠的蛋白导致异常的细胞功能。 这项建议研究蛋白酶如何展开蛋白质。生物学已经达到了一个阶段，可以确定细胞中过程的详细机制。 为了让学生为实验生物学的新时代做好准备，将开发一门关于细胞生物学定量方法的实验室课程。