Jin Mingliang, Zang Yunxiang, Wang Huping, Cong Yao
Key Laboratory of RNA Innovation, Science and Engineering, Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, China.
The Research Center of Chiral Drugs, Innovation Research Institute of Traditional Chinese Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai, China.
QRB Discov. 2024 Dec 16;6:e7. doi: 10.1017/qrd.2024.17. eCollection 2025.
The TRiC/CCT complex assists in the folding of approximately 10% of cytosolic proteins through an ATP-driven conformational cycle, playing a crucial role in maintaining protein homeostasis. Despite our understanding of ATP-driven TRiC ring closing and substrate folding, the process and mechanisms underlying TRiC ring-opening and substrate release remain largely unexplored. In this study, by determining an ensemble of cryo-EM structures of yeast TRiC in the presence of ADP, including three intermediate transition states, we present a comprehensive picture of the TRiC ring-opening process. During this process, CCT3 detects the loss of γ-phosphate and initiates with the dynamics of its apical protrusion, and expands to the outward leaning of the consecutive CCT6/8/7/5 subunits. This is followed by significant movements of CCT2, CCT4, and especially CCT1 subunits, resulting in the opening of the TRiC rings. We also observed an unforeseen temporary separation between the two rings in the CCT2 side, coordinating the release of the originally locked CCT4 N-terminus, which potentially participates in the ring-opening process. Collectively, our study reveals a stepwise TRiC ring-opening mechanism, provides a comprehensive view of the TRiC conformational landscape, and sheds lights on its subunit specificity in sensing nucleotide status and substrate release. Our findings deepen our understanding of protein folding assisted by TRiC and may inspire new strategies for the diagnosis and treatment of related diseases.
TRiC/CCT复合物通过ATP驱动的构象循环协助大约10%的胞质蛋白折叠，在维持蛋白质稳态中发挥关键作用。尽管我们了解ATP驱动的TRiC环闭合和底物折叠过程，但TRiC环打开和底物释放的过程及机制在很大程度上仍未得到探索。在本研究中，通过确定存在ADP时酵母TRiC的一系列冷冻电镜结构，包括三个中间过渡态，我们呈现了TRiC环打开过程的全貌。在此过程中，CCT3检测到γ-磷酸的丢失并以其顶端突起的动态变化开始，随后连续的CCT6/8/7/5亚基向外倾斜。接着，CCT2、CCT4，尤其是CCT1亚基发生显著移动，导致TRiC环打开。我们还观察到CCT2侧的两个环之间出现了意外的暂时分离，这协调了原本锁定的CCT4 N端的释放，其可能参与了环打开过程。总体而言，我们的研究揭示了TRiC环打开的逐步机制，提供了TRiC构象景观的全貌，并阐明了其在感知核苷酸状态和底物释放方面的亚基特异性。我们的发现加深了我们对TRiC协助蛋白质折叠的理解，并可能为相关疾病的诊断和治疗激发新的策略。
