Ehrnsperger M, Lilie H, Gaestel M, Buchner J
Institut für Biophysik und Physikalische Biochemie, Universität Regensburg, 93040 Regensburg, Germany.
J Biol Chem. 1999 May 21;274(21):14867-74. doi: 10.1074/jbc.274.21.14867.
Small heat shock proteins (sHsps), including alpha-crystallin, represent a conserved and ubiquitous family of proteins. They form large oligomers, ranging in size from 140 to more than 800 kDa, which seem to be important for the interaction with non-native proteins as molecular chaperones. Here we analyzed the stability and oligomeric structure of murine Hsp25 and its correlation with function. Upon unfolding, the tertiary and quaternary structure of Hsp25 is rapidly lost, whereas the secondary structure remains remarkably stable. Unfolding is completely reversible, leading to native hexadecameric structures. These oligomers are in a concentration-dependent equilibrium with tetramers and dimers, indicating that tetramers assembled from dimers represent the basic building blocks of Hsp25 oligomers. At high temperatures, the Hsp25 complexes increase in molecular mass, consistent with the appearance of "heat shock granules" in vivo after heat treatment. This high molecular mass "heat shock form" of Hsp25 is in a slow equilibrium with hexadecameric Hsp25. Thus, it does not represent an off-pathway reaction. Interestingly, the heat shock form exhibits unchanged chaperone activity even after incubation at 80 degrees C. We conclude that Hsp25 is a dynamic tetramer of tetramers with a unique ability to refold and reassemble into its active quaternary structure after denaturation. So-called heat shock granules, which have been reported to appear in response to stress, seem to represent a novel functional species of Hsp25.
小热休克蛋白（sHsps），包括α-晶状体蛋白，是一类保守且普遍存在的蛋白质家族。它们形成大小从140 kDa到超过800 kDa不等的大寡聚体，这些寡聚体作为分子伴侣与非天然蛋白质相互作用似乎很重要。在此，我们分析了小鼠Hsp25的稳定性和寡聚结构及其与功能的相关性。在去折叠过程中，Hsp25的三级和四级结构迅速丧失，而二级结构仍保持显著稳定。去折叠是完全可逆的，会形成天然的十六聚体结构。这些寡聚体与四聚体和二聚体处于浓度依赖性平衡，表明由二聚体组装而成的四聚体是Hsp25寡聚体的基本构建单元。在高温下，Hsp25复合物的分子量增加，这与热处理后体内出现的“热休克颗粒”一致。这种高分子量的Hsp25“热休克形式”与十六聚体Hsp25处于缓慢平衡状态。因此，它不代表一条非主要反应途径。有趣的是，即使在80℃孵育后，热休克形式仍表现出不变的伴侣活性。我们得出结论，Hsp25是一种由四聚体组成的动态四聚体，具有在变性后重新折叠并重新组装成其活性四级结构的独特能力。据报道，对应激作出反应而出现的所谓热休克颗粒似乎代表了Hsp25的一种新型功能形式。
