Co-chaperone regulation of conformational switching in the Hsp90 ATPase cycle

Siligardi, Giuliano, Hu, Bin, Panaretou, Barry, Piper, Peter W, Pearl, Laurence H and Prodromou, Chrisostomos (2004) Co-chaperone regulation of conformational switching in the Hsp90 ATPase cycle. Journal of Biological Chemistry, 279 (50). pp. 51989-51998. ISSN 0021-9258

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ATP hydrolysis by the Hsp90 molecular chaperone requires a connected set of conformational switches triggered by ATP binding to the N-terminal domain in the Hsp90 dimer. Central to this is a segment of the structure, which closes like a "lid" over bound ATP, promoting N-terminal dimerization and assembly of a competent active site. Hsp90 mutants that influence these conformational switches have strong effects on ATPase activity. ATPase activity is specifically regulated by Hsp90 co-chaperones, which directly influence the conformational switches. Here we have analyzed the effect of Hsp90 mutations on binding (using isothermal titration calorimetry and difference circular dichroism) and ATPase regulation by the co-chaperones Aha1, Sti1 (Hop), and Sba1 (p23). The ability of Sti1 to bind Hsp90 and arrest its ATPase activity was not affected by any of the mutants screened. Sba1 bound in the presence of AMPPNP to wild-type and ATPase hyperactive mutants with similar affinity but only very weakly to hypoactive mutants despite their wild-type ATP affinity. Unexpectedly, in all cases Sba1 bound to Hsp90 with a 1:2 molar stoichiometry. Aha1 binding to mutants was similar to wild-type, but the -fold activation of their ATPase varied substantially between mutants. Analysis of complex formation with co-chaperone mixtures showed Aha1 and p50cdc37 able to bind Hsp90 simultaneously but without direct interaction. Sba1 and p50cdc37 bound independently to Hsp90-AMPPNP but not together. These data indicated that Sba1 and Aha1 regulate Hsp90 by influencing the conformational state of the "ATP lid" and consequent N-terminal dimerization, whereas Sti1 does not.

Item Type: Article
Keywords: Adenosine Triphosphatases/*chemistry/genetics/*metabolism Adenosine Triphosphate/metabolism Adenylyl Imidodiphosphate/metabolism Calorimetry Chaperonins Dimerization Fungal Proteins/genetics/metabolism HSP90 Heat-Shock Proteins/*chemistry/genetics/*metabolism Heat-Shock Proteins Hydrolysis Models, Molecular Molecular Chaperones/genetics/metabolism Mutation Protein Conformation Protein Structure, Quaternary Recombinant Proteins/chemistry/genetics/metabolism Saccharomyces cerevisiae/genetics/metabolism Saccharomyces cerevisiae Proteins/*chemistry/genetics/*metabolism
Schools and Departments: School of Life Sciences > Biochemistry
Subjects: Q Science > QD Chemistry > QD0241 Organic chemistry > QD0415 Biochemistry
Depositing User: Chrisostomos Prodromou
Date Deposited: 25 Feb 2015 10:22
Last Modified: 25 Feb 2015 10:22
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