Borges Júlio C, Seraphim Thiago V, Dores-Silva Paulo R, Barbosa Leandro R S
Instituto de Química de São Carlos, Universidade de São Paulo, São Carlos, SP, Brazil.
Instituto de Física, Universidade de São Paulo, São Paulo, SP, Brazil.
Biophys Rev. 2016 Jun;8(2):107-120. doi: 10.1007/s12551-016-0194-x. Epub 2016 Mar 4.
Intrinsic flexibility is closely related to protein function, and a plethora of important regulatory proteins have been found to be flexible, multi-domain or even intrinsically disordered. On the one hand, understanding such systems depends on how these proteins behave in solution. On the other, small-angle X-ray scattering (SAXS) is a technique that fulfills the requirements to study protein structure and dynamics relatively quickly with few experimental limitations. Molecular chaperones from Hsp70 and Hsp90 families are multi-domain proteins containing flexible and/or disordered regions that play central roles in cellular proteostasis. Here, we review the structure and function of these proteins by SAXS. Our general approach includes the use of SAXS data to determine size and shape parameters, as well as protein shape reconstruction and their validation by using accessory biophysical tools. Some remarkable examples are presented that exemplify the potential of the SAXS technique. Protein structure can be determined in solution even at limiting protein concentrations (for example, human mortalin, a mitochondrial Hsp70 chaperone). The protein organization, flexibility and function (for example, the J-protein co-chaperones), oligomeric status, domain organization, and flexibility (for the Hsp90 chaperone and the Hip and Hep1 co-chaperones) may also be determined. Lastly, the shape, structural conservation, and protein dynamics (for the Hsp90 chaperone and both p23 and Aha1 co-chaperones) may be studied by SAXS. We believe this review will enhance the application of the SAXS technique to the study of the molecular chaperones.
内在柔韧性与蛋白质功能密切相关，并且已发现大量重要的调节蛋白具有柔韧性、多结构域甚至内在无序。一方面，理解这类系统取决于这些蛋白质在溶液中的行为方式。另一方面，小角X射线散射（SAXS）是一种能够相对快速地研究蛋白质结构和动力学且实验限制较少的技术。来自Hsp70和Hsp90家族的分子伴侣是包含柔性和/或无序区域的多结构域蛋白质，它们在细胞蛋白质稳态中起核心作用。在此，我们通过SAXS综述这些蛋白质的结构和功能。我们的一般方法包括利用SAXS数据确定大小和形状参数，以及进行蛋白质形状重建并通过使用辅助生物物理工具对其进行验证。文中给出了一些显著的例子，例证了SAXS技术的潜力。即使在蛋白质浓度有限的情况下（例如，人线粒体Hsp70伴侣蛋白mortalin），也能在溶液中确定蛋白质结构。还可以确定蛋白质的组织形式、柔韧性和功能（例如，J蛋白共伴侣）、寡聚状态、结构域组织以及柔韧性（对于Hsp90伴侣蛋白以及Hip和Hep1共伴侣）。最后，可以通过SAXS研究形状、结构保守性以及蛋白质动力学（对于Hsp90伴侣蛋白以及p23和Aha1共伴侣）。我们相信这篇综述将促进SAXS技术在分子伴侣研究中的应用。
