Heterogeneity and dynamics in the assembly of the Heat Shock Protein 90 chaperone complexes
Heterogeneity and dynamics in the assembly of the Heat Shock Protein 90 chaperone complexes
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DOI：
10.1073/pnas.1106261108
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发表时间：
2011-11-01
影响因子：
11.1
通讯作者：
Robinson, Carol V.
中科院分区：
文献类型：
--
作者：
Ebong, Ima-obong;Morgner, Nina;Robinson, Carol V.
Hsp90周期依赖于一系列伴侣蛋白的协同活性，包括Hop、Hsp70和肽基脯氨酸异构酶（如FKBP52）。使用质谱法，我们研究了这些辅伴侣的添加顺序及其对所得到的含hsp90配合物的化学计量学和组成的影响。我们的研究结果表明，单分子Hop特异性结合Hsp90二聚体，而FKBP52结合Hsp90的单分子和二聚体形式。通过与Hop和FKBP52，或与FKBP52和Hop形成Hsp90配合物，我们监测了含有两种伴体的主要不对称三联物的形成。这种不对称复合物随后能够与伴侣蛋白Hsp70相互作用，形成包含所有四种蛋白质的四级复合物。监测这些复合物在形成过程中和平衡状态下的种群，使我们能够对复杂的地层进行建模，并提取14种不同的K-D值。在相同的缓冲条件下，用相同的方法从一个复杂的系统、从8个不同的数据集同时计算K(D)s，可以得到一组自一致的值。在这种情况下，K-D值提供了对10个含Hsp90复合物组装的见解，并为Hsp90伴侣蛋白周期中中间产物的细胞异质性和普遍性提供了理论依据。
The Hsp90 cycle depends on the coordinated activity of a range of cochaperones, including Hop, Hsp70 and peptidyl-prolyl isomerases such as FKBP52. Using mass spectrometry, we investigate the order of addition of these cochaperones and their effects on the stoichiometry and composition of the resulting Hsp90-containing complexes. Our results show that monomeric Hop binds specifically to the Hsp90 dimer whereas FKBP52 binds to both monomeric and dimeric forms of Hsp90. By preforming Hsp90 complexes with either Hop, followed by addition of FKBP52, or with FKBP52 and subsequent addition of Hop, we monitor the formation of a predominant asymmetric ternary complex containing both cochaperones. This asymmetric complex is subsequently able to interact with the chaperone Hsp70 to form quaternary complexes containing all four proteins. Monitoring the population of these complexes during their formation and at equilibrium allows us to model the complex formation and to extract 14 different K-D values. This simultaneous calculation of the K(D)s from a complex system with the same method, from eight deferent datasets under the same buffer conditions delivers a self-consistent set of values. In this case, the K-D values afford insights into the assembly of ten Hsp90-containing complexes and provide a rationale for the cellular heterogeneity and prevalence of intermediates in the Hsp90 chaperone cycle.