Refolding activity of bacterial Hsp90 in vivo reveals ancient chaperoning function

The conserved molecular chaperones Hsp70 and Hsp90 play a key role in protein folding. Mechanistically, Hsp90 acts downstream from Hsp70 solving an Hsp70-inflicted folding block. It is unclear, though, when and to which extend the concerted action of this cascade becomes crucial in living organisms. Here we show that, in E. coli cells, Hsp90 dramatically improves protein refolding after heat stress while it is dispensable for de novo folding. We found that Hsp90 inhibition effectively reduced the refolding yields in vivo, leading to strongly reduced enzymatic activity of the paradigmatic chaperone client luciferase and broadly increased aggregation of the E. coli proteome. Additionally, the presence of Hsp90 during refolding reduces the net ATP consumption presumably by sparing the substrate binding-and-release cycles on Hsp70. This mechanism explains how the cooperation of Hsp90 with the Hsp70 chaperone system creates robust folding machinery in a sustainable manner. Together, we describe a general function for bacterial Hsp90 as a key factor of the folding cascade, which may be the ancient activity of this evolutionary conserved machine.