Periplasmic chaperones Skp and SurA are essential players in outer membrane protein (OMP) biogenesis. They prevent unfolded OMPs from misfolding during their passage through the periplasmic space and aid in the disassembly of OMP aggregates under cellular stress conditions. However, functionally important links between interaction mechanisms, structural dynamics, and energetics that underpin both Skp and SurA association with OMPs have remained largely unresolved. Here, using single-molecule fluorescence spectroscopy, we dissect the conformational dynamics and thermodynamics of Skp and SurA binding to unfolded OmpX, and explore their disaggregase activities. We show that both chaperones expand unfolded OmpX distinctly and induce microsecond chain reconfigurations in the client OMP structure. We further reveal that Skp and SurA bind their substrate in a fine-tuned thermodynamic process via enthalpy-entropy compensation. Finally, we observed synergistic activity of both chaperones in the disaggregation of oligomeric OmpX aggregates. Our findings provide an intimate view into the multi-faceted functionalities of Skp and SurA and the fine-tuned balance between conformational flexibility and underlying energetics in aiding chaperone action during OMP biogenesis.

中文翻译：

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伴侣Skp和SurA动态扩展未折叠的外膜蛋白X并协同拆卸寡聚体

周质伴侣Skp和SurA是外膜蛋白（OMP）生物发生中必不可少的参与者。它们可防止未折叠的OMP在穿过周质空间的过程中错误折叠，并有助于在细胞应激条件下分解OMP聚集体。但是，相互作用机制，结构动力学和能量学之间的功能上重要的联系（很大程度上支持Skp和SurA与OMP的联系）仍未解决。在这里，使用单分子荧光光谱，我们分析了Skp和SurA与未折叠的OmpX结合的构象动力学和热力学，并探讨了它们的disagggregase活性。我们显示这两个伴侣明显扩展展开的OmpX，并在客户端OMP结构中诱导微秒链重新配置。我们进一步揭示，Skp和SurA通过焓-熵补偿在微调的热力学过程中结合其底物。最后，我们观察到两种分子伴侣在寡聚OmpX聚集体分解中的协同活性。我们的发现为Skp和SurA的多方面功能以及构象柔韧性和潜在的能量学之间的微调平衡提供了亲密的视角，以帮助OMP生物合成过程中的伴侣作用。