title: Specificity in substrate and cofactor recognition by the N-terminal domain of the chaperone ClpX.
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.
authors.affiliation: One King's College Circle, Medical Sciences Building, Department of Biochemistry, University of Toronto, Toronto, ON, Canada M5S 1A8.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Thibault
authors.forename: Jovana
authors.initials: J
authors.lastname: Yudin
authors.forename: Philip
authors.initials: P
authors.lastname: Wong
authors.forename: Vladimir
authors.initials: V
authors.lastname: Tsitrin
authors.forename: Remco
authors.initials: R
authors.lastname: Sprangers
authors.forename: Rongmin
authors.initials: R
authors.lastname: Zhao
authors.forename: Walid A
authors.initials: WA
authors.lastname: Houry
mesh_terms: D000074183:ATPases Associated with Diverse Cellular Activities; D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003067:Coenzymes; D004252:DNA Mutational Analysis; D049071:Endopeptidase Clp / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity
chemical_list: D002352:Carrier Proteins; D003067:Coenzymes; D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; C477700:sspB protein, E coli; D049071:Endopeptidase Clp; D000251:Adenosine Triphosphatases; C088072:ClpX protein, E coli; D000074183:ATPases Associated with Diverse Cellular Activities