Wojtyra Urszula A, Thibault Guillaume, Tuite Ashleigh, Houry Walid A
Department of Biochemistry, Medical Sciences Building, 1 King's College Circle, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
J Biol Chem. 2003 Dec 5;278(49):48981-90. doi: 10.1074/jbc.M307825200. Epub 2003 Aug 23.
Clp ATPases are unique chaperones that promote protein unfolding and subsequent degradation by proteases. The mechanism by which this occurs is poorly understood. Here we demonstrate that the N-terminal domain of ClpX is a C4-type zinc binding domain (ZBD) involved in substrate recognition. ZBD forms a very stable dimer that is essential for promoting the degradation of some typical ClpXP substrates such as lambdaO and MuA but not GFP-SsrA. Furthermore, experiments indicate that ZBD contains a primary binding site for the lambdaO substrate and for the cofactor SspB. Removal of ZBD from the ClpX sequence renders the ATPase activity of ClpX largely insensitive to the presence of ClpP, substrates, or the SspB cofactor. All these results indicate that ZBD plays an important role in the ClpX mechanism of function and that ATP binding and/or hydrolysis drives a conformational change in ClpX involving ZBD.
Clp ATP酶是一类独特的伴侣蛋白，可促进蛋白质解折叠并随后由蛋白酶进行降解。其发生机制目前尚不清楚。在此我们证明，ClpX的N端结构域是一个参与底物识别的C4型锌结合结构域（ZBD）。ZBD形成一个非常稳定的二聚体，这对于促进某些典型的ClpXP底物（如λO和MuA，但不包括GFP-SsrA）的降解至关重要。此外，实验表明ZBD含有λO底物和辅因子SspB的主要结合位点。从ClpX序列中去除ZBD会使ClpX的ATP酶活性在很大程度上对ClpP、底物或SspB辅因子的存在不敏感。所有这些结果表明，ZBD在ClpX的功能机制中起重要作用，并且ATP结合和/或水解驱动了涉及ZBD的ClpX构象变化。
