Molecular chaperones are proteins that assist the folding, unfolding, and 
remodeling of other proteins. In eukaryotes, heat shock protein 90 (Hsp90) 
proteins are essential ATP-dependent molecular chaperones that remodel and 
activate hundreds of client proteins with the assistance of cochaperones. In 
Escherichia coli, the activity of the Hsp90 homolog, HtpG, has remained elusive. 
To explore the mechanism of action of E. coli Hsp90, we used in vitro protein 
reactivation assays. We found that E. coli Hsp90 promotes reactivation of 
heat-inactivated luciferase in a reaction that requires the prokaryotic Hsp70 
chaperone system, known as the DnaK system. An Hsp90 ATPase inhibitor, 
geldanamycin, inhibits luciferase reactivation demonstrating the importance of 
the ATP-dependent chaperone activity of E. coli Hsp90 during client protein 
remodeling. Reactivation also depends upon the ATP-dependent chaperone activity 
of the DnaK system. Our results suggest that the DnaK system acts first on the 
client protein, and then E. coli Hsp90 and the DnaK system collaborate 
synergistically to complete remodeling of the client protein. Results indicate 
that E. coli Hsp90 and DnaK interact in vivo and in vitro, providing additional 
evidence to suggest that E. coli Hsp90 and the DnaK system function together.