THE ALLOSTERIC MECHANISM OF HSP70

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 分子伴侣通过保护未折叠和错误折叠的蛋白质，直到它们能够折叠或组装成复合体，从而帮助促进成功的折叠和组装，在细胞维持和应对压力方面发挥着至关重要的作用。他们还与降解机械合作，管理细胞的“质量控制”，防止潜在有害的错误折叠产品的积累。伴侣蛋白共同作用形成一个复杂的蛋白质平衡网络，这对细胞的生存至关重要。无处不在的Hsp70蛋白家族的伴侣蛋白防止未折叠蛋白质中的错误相互作用，这些相互作用可能导致错误折叠或聚集，这些错误相互作用是由ATP水解酶调节的客户蛋白的结合和释放，并进一步受到辅助伴侣蛋白的调节，如核苷酸交换因子(NEF)和Hsp40s。了解Hsp70的细胞作用需要深入阐明Hsp70的变构机制以及与辅助伴侣的相互作用；这对于开发基于Hsp70伴侣调节的治疗策略是至关重要的。我们在Acert设施的工作目标是获得关键信息，以全面描述大肠杆菌Hsp70，DNAK在其整个功能变构循环中以及在与辅助伴侣相互作用时的结构系综。我们将确定在整个蛋白质中具有战略意义的各种自旋对之间的距离分布。这些距离分布将提供关于DNAK在各种核苷酸或底物或共伴侣结合状态下填充的结构的信息。此外，我们将使用双电子-电子共振(DER)实验来识别这些集合中稀疏分布的构象。这些轮廓，结合在美国马萨诸塞州大学收集的顺磁弛豫增强核磁共振数据，将使我们能够绘制出HSP70的变构景观。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Molecular chaperones play essential roles in cellular maintenance and response to stress by protecting unfolded and misfolded proteins until they can be folded or assembled into complexes and then helping promote successful folding and assembly. They also partner with degradation machinery to manage cellular 'quality control' and prevent accumulation of potentially deleterious misfolded products. Chaperones work together to form a complex proteostasis network that is essential for cell survival. The ubiquitous Hsp70 protein family of chaperones prevents incorrect interactions in unfolded proteins that can lead to misfolding or aggregation by cycles of binding and release of client proteins regulated by ATP hydrolysis and further modulated by co-chaperones such as nucleotide exchange factors (NEFs) and Hsp40s. Understanding cellular roles of Hsp70s requires in-depth elucidation of the Hsp70 allosteric mechanism and interactions with co-chaperones; this is essential in order to develop therapeutic strategies based on modulation of Hsp70 chaperones. The goal of our work at the ACERT facility is to obtain crucial information for a full description of the structural ensembles of the E. coli Hsp70, DnaK, throughout its functional allosteric cycle and upon interaction with co-chaperones. We will determine distance profiles between various spin pairs strategically located throughout the protein. These distance profiles will provide information about the structures populated by DnaK in various nucleotide- or substrate- or co-chaperone-bound states. In addition, we will use double electron-electron resonance (DEER) experiments to identify sparsely populated conformers within these ensembles. These profiles, in conjunction with paramagnetic relaxation enhancement NMR data being collected at U. Mass will enable us to map out the allosteric landscape of an Hsp70.