Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520, USA.
Protein Sci. 2010 Jan;19(1):19-25. doi: 10.1002/pro.278.
The co-chaperone Hsp70-Hsp90 organizing protein (HOP) plays a central role in protein folding in vivo, binding to both Hsp70 and Hsp90 and bringing them together in a functional complex. Reports in the literature concerning the oligomeric state of HOP have been inconsistent-is it a monomer, dimer, or higher order oligomer? Knowing the oligomeric state of HOP is important, because it places limits on the number and types of multiprotein complexes that can form during the folding cycle. Thus, the number of feasible models is simplified. Here, we explicitly investigate the oligomeric state of HOP using three complementary methods: gel filtration chromatography, sedimentation equilibrium analytical ultracentrifugation (AUC), and an in vivo coexpression assay. We find that HOP does not behave like a monomeric globular protein on gel filtration. Rather its behavior is consistent with it being either an elongated monomer or a dimer. We follow-up on these studies using sedimentation equilibrium AUC, which separates on the basis of molecular weight (MW), independent of shape. Sedimentation equilibrium AUC clearly shows that HOP is a monomer, with no indication of higher MW species. Finally, we use an in vivo coexpression assay that also supports the conclusion that HOP is a monomer.
共伴侣热休克蛋白 70-90 组织蛋白（HOP）在体内蛋白质折叠中发挥核心作用，与热休克蛋白 70 和热休克蛋白 90 结合，并将它们结合在一个功能复合物中。文献中关于 HOP 寡聚状态的报告一直不一致——它是单体、二聚体还是更高阶的寡聚体？了解 HOP 的寡聚状态很重要，因为它限制了在折叠循环中可以形成的多蛋白复合物的数量和类型。因此，可行的模型数量被简化。在这里，我们使用三种互补的方法来明确研究 HOP 的寡聚状态：凝胶过滤色谱、沉降平衡分析超速离心（AUC）和体内共表达测定。我们发现 HOP 在凝胶过滤上的行为不像单体球状蛋白。相反，它的行为与其是伸长的单体或二聚体一致。我们使用沉降平衡 AUC 跟进这些研究，沉降平衡 AUC 基于分子量（MW）分离，与形状无关。沉降平衡 AUC 清楚地表明 HOP 是单体，没有更高 MW 物种的迹象。最后，我们使用体内共表达测定，也支持 HOP 是单体的结论。
