Zentrum für Molekulare Biologie der Universität Heidelberg (ZMBH), Heidelberg, Germany.
Nat Struct Mol Biol. 2012 Dec;19(12):1347-55. doi: 10.1038/nsmb.2442. Epub 2012 Nov 18.
Bacteria, fungi and plants rescue aggregated proteins using a powerful bichaperone system composed of an Hsp70 chaperone and an Hsp100 AAA+ disaggregase. In Escherichia coli, the Hsp70 chaperone DnaK binds aggregates and targets the disaggregase ClpB to the substrate. ClpB hexamers use ATP to thread substrate polypeptides through the central pore, driving disaggregation. How ClpB finds DnaK and regulates threading remains unclear. To dissect the disaggregation mechanism, we separated these steps using primarily chimeric ClpB-ClpV constructs that directly recognize alternative substrates, thereby obviating DnaK involvement. We show that ClpB has low intrinsic disaggregation activity that is normally repressed by the ClpB middle (M) domain. In the presence of aggregate, DnaK directly binds M-domain motif 2, increasing ClpB ATPase activity to unleash high ClpB threading power. Our results uncover a new function for Hsp70: the coupling of substrate targeting to AAA+ chaperone activation at aggregate surfaces.
细菌、真菌和植物使用一种由 Hsp70 伴侣蛋白和 Hsp100 AAA+ 解聚酶组成的强大双伴侣系统来拯救聚集的蛋白质。在大肠杆菌中，Hsp70 伴侣蛋白 DnaK 结合聚集体，并将解聚酶 ClpB 靶向底物。ClpB 六聚体利用 ATP 将底物多肽穿过中央孔，从而驱动解聚。ClpB 如何找到 DnaK 并调节穿线仍然不清楚。为了剖析解聚机制，我们主要使用嵌合 ClpB-ClpV 构建体分离这些步骤，这些构建体可以直接识别替代底物，从而避免 DnaK 的参与。我们表明，ClpB 具有低的固有解聚活性，通常被 ClpB 中间 (M) 结构域抑制。在聚集体存在的情况下，DnaK 直接结合 M 结构域基序 2，增加 ClpB ATPase 活性，释放高 ClpB 穿线能力。我们的结果揭示了 Hsp70 的一个新功能：将底物靶向与在聚集体表面激活 AAA+ 伴侣蛋白耦合。
