Thibault Guillaume, Yudin Jovana, Wong Philip, Tsitrin Vladimir, Sprangers Remco, Zhao Rongmin, Houry Walid A
One King's College Circle, Medical Sciences Building, Department of Biochemistry, University of Toronto, Toronto, ON, Canada M5S 1A8.
Proc Natl Acad Sci U S A. 2006 Nov 21;103(47):17724-9. doi: 10.1073/pnas.0601505103. Epub 2006 Nov 7.
Clp ATPases are a unique group of ATP-dependent chaperones supporting targeted protein unfolding and degradation in concert with their respective proteases. ClpX is a representative member of these ATPases; it consists of two domains, a zinc-binding domain (ZBD) that forms dimers and a AAA+ ATP-binding domain that arranges into a hexamer. Analysis of the binding preferences of these two domains in ClpX revealed that both domains preferentially bind to hydrophobic residues but have different sequence preferences, with the AAA+ domain preferentially recognizing a wider range of specific sequences than ZBD. As part of this analysis, the binding site of the ClpX dimeric cofactor, SspB2, on ZBD in ClpX was determined by NMR and mutational analysis. The SspB C terminus was found to interact with a hydrophobic patch on the surface of ZBD. The affinity of SspB2 toward ZBD2 and the geometry of the SspB2-ZBD2 complex were investigated by using the newly developed quantitative optical biosensor method of dual polarization interferometry. The data suggest a model for the interaction between SspB2 and the ClpX hexamer.
Clp ATP酶是一类独特的依赖ATP的伴侣蛋白，它们与各自的蛋白酶协同作用，支持靶向蛋白的解折叠和降解。ClpX是这些ATP酶的一个代表性成员；它由两个结构域组成，一个形成二聚体的锌结合结构域（ZBD）和一个排列成六聚体的AAA+ ATP结合结构域。对ClpX中这两个结构域的结合偏好性分析表明，这两个结构域都优先结合疏水残基，但具有不同的序列偏好，其中AAA+结构域比ZBD优先识别更广泛的特定序列。作为该分析的一部分，通过核磁共振和突变分析确定了ClpX二聚体辅因子SspB2在ClpX的ZBD上的结合位点。发现SspB的C末端与ZBD表面的一个疏水区域相互作用。通过使用新开发的双偏振干涉定量光学生物传感器方法，研究了SspB2对ZBD2的亲和力以及SspB2-ZBD2复合物的几何结构。这些数据提出了一个SspB2与ClpX六聚体相互作用的模型。
