Structure and function of ClpXP

ClpXP的结构和功能

• 批准号: 10198307
• 负责人: Robert T Sauer
• 金额: $ 38.57万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10198307
• 关键词: ATP Hydrolysis; ATP phosphohydrolase; ATP-Dependent Proteases; Address; Antibiotics; Basic Science; Biochemical; Biotechnology; C-terminal; Cell physiology; Cells; Coupled; Defect; Disease; Ensure; Enzymes; Escherichia coli; Family; Goals; Growth; Health Promotion; Infertility; Light; Mammals; Mechanics; Medicine; Mitochondria; Molecular; Mycobacterium tuberculosis; N-terminal; Pathogenesis; Peptide Hydrolases; Play; Positioning Attribute; Proteins; Role; Specificity; Structure; Time; Work; experimental study; hearing impairment; heme biosynthesis; polypeptide; prevent; protein degradation; protein folding

Project Summary AAA+ proteases remove toxic proteins and regulate many other important cellular processes that promote health and prevent disease. At the same time, protein degradation must be carefully regulated. AAA+ proteases assemble into multi-subunit structures with an internal proteolytic chamber, accessible through narrow channels that exclude natively folded proteins. This mechanism protects most proteins from unintended degradation and requires specific substrates to be recognized, unfolded, and then translocated into the degradation chamber. In the AAA+ ClpXP protease, for example, a ring hexamer of ClpX uses the energy of ATP hydrolysis to unfold specific target proteins and translocate them into ClpP for degradation. ClpXP is one of the best-characterized AAA+ proteases and is a paradigm for other ATP- dependent proteases and AAA+ remodeling machines. These ATP-fueled enzymes perform a wide variety of mechanical remodeling, transport, and regulatory tasks in the cell. In mammals, loss of mitochondrial ClpP results in infertility, hearing loss, and growth defects, whereas mitochondrial ClpX plays an important role in heme biosynthesis. Bacterial ClpXP can promote pathogenesis and is a validated antibiotic target in M. tuberculosis. Substantial progress has been made in understanding the general biochemical and structural features of E. coli ClpXP and other AAA+ enzymes but important and fundamental questions concerning the molecular mechanisms of these machines remain. For example, it is not known how ClpX identifies many classes of N-terminal and C-terminal degrons, whether ClpX rotates with respect to ClpP during normal function, whether proofreading helps ensure degradation specificity, how multiple substrate chains can be simultaneously translocated through the ClpX channel, what the detailed and interacts with polypeptide substrates during mechanical unfolding, whether ATP hydrolysis can occur at multiple positions in the spiral ClpX ring or only at one or a few special positions, and how the detailed ATPase cycle is coupled to mechanical work. The experiments described in this proposal will address these questions and provide a conceptual framework applicable to studies of the entire superfamily of AAA+ machines.

项目摘要 AAA+蛋白酶去除有毒蛋白并调节许多其他重要的细胞过程 促进健康并预防疾病。同时，蛋白质降解必须是 精心监管。 AAA+蛋白酶与内部组合成组合 蛋白水解室，可通过排除本内折叠蛋白的狭窄通道访问。 这种机制可保护大多数蛋白质免受意外降解的侵害，并需要特定 底物被识别，展开，然后易位到降解室中。在 例如，AAA+ CLPXP蛋白酶，例如，CLPX的六聚体使用ATP的能量 水解以展开特定的靶蛋白并将其转移到CLPP中以降解。 CLPXP是特征最佳的AAA+蛋白酶之一，是其他ATP-的范式 依赖蛋白酶和AAA+重塑机。这些ATP燃料酶执行 细胞中的各种机械重塑，运输和调节任务。在哺乳动物中， 线粒体CLPP的丧失会导致不育，听力损失和生长缺陷，而 线粒体CLPX在血红素生物合成中起重要作用。细菌CLPXP可以促进 发病机理，是结核分枝杆菌中经过验证的抗生素靶标。实质性进展已有 是在理解大肠杆菌Clpxp的一般生化和结构特征方面制作的 和其他AAA+酶，但有关分子的重要和基本问题 这些机器的机制仍然存在。例如，尚不知道CLPX如何识别许多 N末端和C末端degrons类别，CLPX是否相对于CLPP旋转 在正常功能期间，是否有助于确保降解特异性，如何 可以通过CLPX通道同时易位多个底物链，什么 机械展开期间的详细并与多肽底物相互作用，是否 ATP水解可以在螺旋CLPX环的多个位置发生，也可以在一个或几个位置发生 特殊位置，以及如何将详细的ATPase周期与机械工作耦合。这 本提案中描述的实验将解决这些问题并提供概念 适用于AAA+机器整个超家族的研究框架。