Hsp90, a crucial molecular chaperone, regulates diverse client proteins, impacting both normal biology and disease. Central to its function is its conformational plasticity, driven by ATPase activity and client interactions. However, comprehensive insights into Hsp90’s dynamic molecular transitions remain elusive. Using solution NMR spectroscopy, we reveal how ATP binding, hydrolysis, and client engagement drive conformational and dynamic shifts in E. coli Hsp90, HtpG, through its chaperone cycle. Pronounced conformational fluctuations occur, especially in regions crucial for nucleotide binding and conformational transitions. ATP binding induces slow-exchanging conformations, representing discrete on-path transition states from open to closed forms, while ATP hydrolysis shifts HtpG into a compact conformation. Client binding acts as an allosteric switch, dynamically priming HtpG for elevated chaperone activity and, therefore, its efficient remodeling. Here, we provide atomic-level insights into Hsp90’s functional mechanism, highlighting the interplay of conformation, dynamics, nucleotide, and client interactions.

Hsp90 是一种重要的分子伴侣，可调节多种客户蛋白，影响正常生物学和疾病。其功能的核心是其构象可塑性，由 ATP 酶活性和客户相互作用驱动。然而，对 Hsp90 动态分子转变的全面了解仍然难以捉摸。使用溶液 NMR 波谱，我们揭示了 ATP 结合、水解和客户参与如何通过其伴侣循环驱动大肠杆菌 Hsp90 （HtpG） 的构象和动态变化。发生明显的构象波动，尤其是在核苷酸结合和构象转换的关键区域。ATP 结合诱导缓慢交换的构象，代表从开放形式到封闭形式的离散路径上转换状态，而 ATP 水解将 HtpG 转变为紧凑的构象。客户端绑定充当变构开关，动态启动 HtpG 以提升伴侣活动，从而有效地重塑。在这里，我们提供了对 Hsp90 功能机制的原子级见解，突出了构象、动力学、核苷酸和客户端相互作用的相互作用。