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Hsp90 from E. coli collaborates with the DnaK chaperone system in client protein remodeling

Wednesday, October 26, 2011 — Poster Session IV

2:00 p.m. – 4:00 p.m.

Natcher Conference Center



* FARE Award Winner


  • O Genest
  • JR Hoskins
  • JL Camberg
  • SM Doyle
  • S Wickner


Molecular chaperones are proteins that assist the folding, unfolding and reactivation of other proteins. In eukaryotes, members of the Hsp90 family are essential ATP-dependent molecular chaperones that, with the assistance of many cochaperones, remodel and activate hundreds of client proteins. Prokaryotic Hsp90 is highly homologous to eukaryotic Hsp90, however, its function has been elusive. To explore the mechanism of E. coli Hsp90 (Hsp90Ec) activity, we developed an in vitro protein remodeling assay using the heat-denatured model substrate luciferase and purified chaperones. We found that reactivation of luciferase requires both Hsp90Ec and the prokaryotic Hsp70 chaperone system, known as the DnaK system and comprised of DnaK and two cochaperones. Using Hsp90Ec mutant proteins defective in ATP hydrolysis and an Hsp90 ATPase inhibitor, geldanamycin, we showed that ATP hydrolysis by Hsp90Ec was essential for protein reactivation in vitro. We also performed in vivo two-hybrid experiments and in vitro protein binding assays that showed Hsp90Ec and DnaK directly interact. These results demonstrate that Hsp90Ec is a bona fide molecular chaperone, able to remodel client proteins in an ATP-dependent reaction. Moreover, it provides a much needed model system to elucidate the ATP-dependent chaperone action of Hsp90 proteins in collaboration with the DnaK/Hsp70 chaperone system.

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