Department of Physics, Informatics and Mathematics, University of Modena and Reggio Emilia, 41125, Modena, Italy.
Institute of Nanoscience S3, Consiglio Nazionale delle Ricerche, 41125, Modena, Italy.
Cell Stress Chaperones. 2020 Jul;25(4):615-628. doi: 10.1007/s12192-020-01096-y. Epub 2020 Apr 6.
Imbalances of cellular proteostasis are linked to ageing and human diseases, including neurodegenerative and neuromuscular diseases. Heat shock proteins (HSPs) and small heat shock proteins (sHSPs) together form a crucial core of the molecular chaperone family that plays a vital role in maintaining cellular proteostasis by shielding client proteins against aggregation and misfolding. sHSPs are thought to act as the first line of defence against protein unfolding/misfolding and have been suggested to act as "sponges" that rapidly sequester these aberrant species for further processing, refolding, or degradation, with the assistance of the HSP70 chaperone system. Understanding how these chaperones work at the molecular level will offer unprecedented insights for their manipulation as therapeutic avenues for the treatment of ageing and human disease. The evolution in single-molecule force spectroscopy techniques, such as optical tweezers (OT) and atomic force microscopy (AFM), over the last few decades have made it possible to explore at the single-molecule level the structural dynamics of HSPs and sHSPs and to examine the key molecular mechanisms underlying their chaperone activities. In this paper, we describe the working principles of OT and AFM and the experimental strategies used to employ these techniques to study molecular chaperones. We then describe the results of some of the most relevant single-molecule manipulation studies on HSPs and sHSPs and discuss how these findings suggest a more complex physiological role for these chaperones than previously assumed.
细胞蛋白质平衡的失衡与衰老和人类疾病有关，包括神经退行性疾病和神经肌肉疾病。热休克蛋白（HSPs）和小分子热休克蛋白（sHSPs）共同构成分子伴侣家族的核心，在维持细胞蛋白质平衡方面发挥着至关重要的作用，它可以保护客户蛋白免受聚集和错误折叠的影响。sHSPs 被认为是对抗蛋白质展开/错误折叠的第一道防线，并且被认为是“海绵”，可以迅速隔离这些异常物种，以便进一步进行处理、重折叠或降解，这需要 HSP70 伴侣系统的协助。了解这些伴侣蛋白在分子水平上的工作机制将为它们的操纵提供前所未有的见解，为治疗衰老和人类疾病提供新的途径。在过去几十年中，单分子力谱技术（如光镊（OT）和原子力显微镜（AFM））的发展使得在单分子水平上探索 HSPs 和 sHSPs 的结构动力学并研究其伴侣活性的关键分子机制成为可能。在本文中，我们描述了 OT 和 AFM 的工作原理以及使用这些技术研究分子伴侣的实验策略。然后，我们描述了一些关于 HSPs 和 sHSPs 的最相关的单分子操纵研究的结果，并讨论了这些发现如何表明这些伴侣蛋白的生理作用比以前假设的更为复杂。
