Principles of substrate recognition by the eukaryotic chaperonin TRiC/CCT

描述(申请人提供)：伴侣蛋白/CCT是真核蛋白质折叠机制的中心组件，估计折叠真核蛋白质组的5%-10%。TIC组装成由两个堆叠的环组成的环状低聚复合体，每个环包含八个不同的亚基。这些环创建了一个中央空腔，在那里未折叠的蛋白质以依赖于ATP的方式结合和折叠。一个基本的问题仍然存在：TIRE如何识别和折叠不同的真核蛋白质子集？我们假设，异源低聚复合体中的每个亚基在底物特异性上都不同。然而，这一假设仍未得到检验，因为人们对电离子-衬底相互作用的性质知之甚少。该建议的目的是确定底物结合位点在TIRE亚基中的位置，并阐明TIRE识别底物的机制。为此，我们将使用多学科方法来比较来自不同TIC底物的基序如何与单个TIC亚基结合。了解底物相互作用将揭示这种伴侣蛋白促进蛋白质折叠的机制。这项工作将对开发新的治疗方法来改善与蛋白质错误折叠相关的疾病具有重要意义。 公共卫生相关性：真核细胞伴侣蛋白trE在许多重要细胞真核蛋白的成熟过程中起着基础性作用，其功能丧失与许多疾病有关，如神经退行性变和癌症。在拟议的实验中，系统地剖析不同底物的伴侣-底物结合作用的结果，将为这些病理的治疗干预提供方向。

DESCRIPTION (provided by applicant): The chaperonin TRiC/CCT is a central component of the eukaryotic protein folding machinery and is estimated to fold 5-10% of the eukaryotic proteome. TRiC assembles into a ring-shaped oligomeric complex that is comprised of two stacked rings each containing eight different subunits. The rings create a central cavity where unfolded proteins bind and fold in an ATP dependent manner. A fundamental question remains: how does TRiC recognize and fold a diverse subset of eukaryotic proteins? We hypothesize that each subunit in the hetero-oligomeric complex differs in substrate specificity. However, this hypothesis remains untested, as little is known about the nature of TRiC-substrate interactions. The goal of this proposal is to identify the location of the substrate binding sites in the TRiC subunits and elucidate the mechanisms underlying substrate recognition by TRiC. To this end, we will use a multidisciplinary approach to compare how motifs derived from different TRiC substrates bind to individual TRiC subunits. Understanding TRiC substrate interactions will reveal the mechanism by which this chaperonin facilitates protein folding. This work will have important implications for the development of novel therapeutic approaches to ameliorate diseases associated with protein misfolding. PUBLIC HEALTH RELEVANCE: The eukaryotic chaperonin TRiC plays a fundamental role in the maturation of many important cellular eukaryotic proteins, and its loss-of-function is associated with numerous diseases such as neurodegeneration and cancer. The results of systematically dissecting the chaperonin-substrate binding interactions for diverse substrates in the proposed experiments, will offer directions for therapeutic interventions in these pathologies.