B CUBE - Center for Molecular Bioengineering, Technische Universität Dresden, 01307 Dresden, Germany.
B CUBE - Center for Molecular Bioengineering, Technische Universität Dresden, 01307 Dresden, Germany;
Proc Natl Acad Sci U S A. 2022 Mar 1;119(9). doi: 10.1073/pnas.2118919119.
Periplasmic chaperones 17-kilodalton protein (Skp) and survival factor A (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 associations 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 multifaceted functionalities of Skp and SurA and the fine-tuned balance between conformational flexibility and underlying energetics in aiding chaperone action during OMP biogenesis.
周质伴侣蛋白 17 千道尔顿蛋白（Skp）和生存因子 A（SurA）是外膜蛋白（OMP）生物发生的重要参与者。它们可以防止未折叠的 OMP 在穿过周质空间时错误折叠，并在细胞应激条件下帮助 OMP 聚集体的解体。然而，支撑 Skp 和 SurA 与 OMP 相互作用的机制、结构动力学和能量学之间的功能重要联系在很大程度上仍未得到解决。在这里，我们使用单分子荧光光谱法，剖析了 Skp 和 SurA 与未折叠的 OmpX 结合的构象动力学和热力学，并研究了它们的解聚酶活性。我们表明，这两种伴侣蛋白都能明显地扩展未折叠的 OmpX，并诱导客户 OMP 结构中的微秒链重新配置。我们进一步揭示 Skp 和 SurA 通过焓熵补偿在精细的热力学过程中结合其底物。最后，我们观察到两种伴侣蛋白在寡聚 OmpX 聚集体的解聚中具有协同活性。我们的发现提供了一个深入了解 Skp 和 SurA 的多方面功能的视角，以及在 OMP 生物发生过程中帮助伴侣蛋白发挥作用时，构象灵活性和潜在能量之间的精细平衡。
