STRUCTURE - FUNCTION AND KINETICS OF AAA+ ATPASES

AAA 腺苷酸酶的结构 - 功能和动力学

• 批准号: 7722747
• 负责人: B TRACY NIXON
• 金额: $ 1.26万
• 依托单位: ILLINOIS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2008-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7722747
• 关键词: ATP Hydrolysis; ATP phosphohydrolase; Address; Apyrase; Binding; Cell division; Cells; Computer Retrieval of Information on Scientific Projects Database; DNA; Data; Defect; Disease; Funding; Gene Expression; Genetic Transcription; Grant; Human; Hydrogen; Hydrolysis; Institution; Kinetics; Mechanics; Membrane; Molecular Machines; Nitrogen Fixation; Organelles; Outcome; Production; Proteins; Research; Research Personnel; Resources; Source; Structure; Time; United States National Institutes of Health; Work; enhancer binding protein; mutant; novel; pathogen; remediation; repaired

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Why do defects in molecular machines (AAA+ ATPases) cause disease? These protein machines convert ATP hydrolysis into mechanical work. Both human cells and disease causing pathogens use this work to physically manipulate proteins or DNA to dismantle and reassemble membranes or other organelles, to replicate DNA and traverse cell division, to repair damaged proteins, or to regulate gene expression. We do not know how these molecular machines convert ATP hydrolysis into mechanical work. Our research focuses on one subset of AAA+ ATPAses, the bacterial-enhancer-binding proteins (EBPs) which use their ATPase activities to regulate transcription of genes needed for harmful activites (diseases, crop damage) or helpful ones (nitrogen fixation, environmental remediation, hydrogen or other metabolite production). In a prior project, we established two mechanisms for regulating the EBP ATPases, and began defining structural changes occuring in their catalytic cycle. The current project completes that objective, also extending it to address the underlying mechanism via structure function studies of mutant forms of ATPase. An unexpected outcome was the discovery of a second form of the ATPase that hydrolyzes ADP to AMP. This novel activity for AAA+ ATPases was discovered because SAXS data showed ADP causing the same conformational changes in a mutant of the ATPase that are caused by ATP binding to the wild type protein. Chromatographic separation of the ATPase from the apyrase form yielded for the first time a crystal structure of the ATP-bound form of this particular AAA+ ATPase, allowing us to further interpret prior SAXS data.

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 为什么分子机器(AAA+ATPase)中的缺陷会导致疾病？这些蛋白质机器将三磷酸腺苷的水解转化为机械功。人类细胞和引起疾病的病原体都利用这种工作来物理地操纵蛋白质或DNA，以拆解和重组膜或其他细胞器，复制DNA并穿越细胞分裂，修复受损的蛋白质，或调节基因表达。我们不知道这些分子机器是如何将三磷酸腺苷的水解转化为机械功的。我们的研究集中在AAA+ATPase的一个子集，细菌增强子结合蛋白(EBPs)，它利用它们的ATPase活性来调节有害活动(疾病、作物损害)或有用活动(固氮、环境修复、氢气或其他代谢物产生)所需基因的转录。 在之前的一个项目中，我们建立了两种调节EBP ATPase的机制，并开始定义在其催化循环中发生的结构变化。目前的项目完成了这一目标，还将其扩展到通过对ATPase突变形式的结构功能研究来解决潜在的机制。一个意想不到的结果是发现了第二种形式的ATPase，它能将ADP水解为AMP。发现AAA+ATPase的这种新活性是因为SAXS数据显示，ADP引起ATPase突变体的构象变化与ATP与野生型蛋白结合所引起的构象变化相同。将ATPase从apyrase形式中分离出来，首次得到了这种特殊的AAA+ATPase的ATP结合形式的晶体结构，使我们能够进一步解释先前的SAXS数据。