NUCLEOTIDE DEPENDENT CONFORMATIONAL CHANGES IN S54-DEPENDENT AAA+ATPASES

S54 依赖的 AAA 腺苷酸酶中核苷酸依赖的构象变化

• 批准号: 7369131
• 负责人: B TRACY NIXON
• 金额: $ 0.44万
• 依托单位: ILLINOIS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-01 至 2007-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7369131
• 关键词: NUCLEOTIDE; DEPENDENT; CONFORMATIONAL; CHANGES; S54

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. It is clear that 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. But, we do not know how these molecular machines convert ATP hydrolysis into mechanical work. A major impediment to defining these mechanisms has been our inability to probe detailed conformational changes that are related to controlling these ATPases as they step through ATP binding, hydrolysis and product release. Our project focused on defining structural changes that occur to regulate assembly of s54-dependent AAA+ ATPases, and that accompany the ATP hydrolysis cycle. These activators remodel the s54-form of bacterial RNA polymerase to enable it to initiate transcription in bacteria. Previous solution structures derived from small angle scattering data (published in 2005) revealed that in one case a regulatory domain adopts two homo-dimeric forms, alternately repressing or derepressing motor assembly by adjacent ATPase domains; in another case, regulatory and ATPase domains were found to cooperate to stabilize the assembled motor. Similar data collected in 2005 revealed a) conformational changes that are coupled with nucleotide status and that mediate two distinct interactions with the target protein, s54, and b) structure of one of those complexes. These latter results are being submitted for publication.

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。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。显然，人类细胞和引起疾病的病原体都利用这一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。但是，我们不知道这些分子机器是如何将ATP水解转化为机械功的。确定这些机制的一个主要障碍是我们无法探索与这些ATPase在ATP结合、水解和产物释放过程中控制相关的详细构象变化。我们的项目重点是定义发生的结构变化，以调节依赖s54的AAA+ATPase的组装，并伴随着ATP水解循环。这些激活剂重塑细菌RNA聚合酶的s54形式，使其能够在细菌中启动转录。根据小角散射数据(2005年发表)得到的先前的溶液结构显示，在一个案例中，调节结构域采用两种同源二聚体形式，通过相邻的ATPase结构域交替抑制或去抑制电机组装；在另一个案例中，调节结构域和ATPase结构域被发现协同稳定组装的电机。2005年收集的类似数据揭示了a)与核苷酸状态相结合的构象变化，以及与目标蛋白s54的两种不同的相互作用，以及b)其中一个复合体的结构。这些后一项研究结果正在提交发表。