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Updated: Jul 19, 2026

Studies of Chaperone-Cochaperone Interactions using Homogenous Bead-Based Assay
Published on: July 21, 2021
Specificity in substrate and cofactor recognition by the N-terminal domain of the chaperone ClpX
Guillaume Thibault1, Jovana Yudin, Philip Wong
1One King's College Circle, Medical Sciences Building, Department of Biochemistry, University of Toronto, Toronto, ON, Canada M5S 1A8.
Abstract:
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.
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