2 CHAPERONE SITES IN HSP90 DIFFERING IN SUBSTRATE-SPECIFICITY AND ATPDEPENDENCE

The abundant molecular chaperone Hsp90 is a key regulator of protein s tructure in the cytosol of eukaryotic cells. Although under physiologi cal conditions a specific subset of proteins is substrate for Hsp90, u nder stress conditions Hsp90 seems to perform more general functions. However, the underlying mechanism of Hsp90 remained enigmatic. Here, w e analyzed the function of conserved Hsp90 domains. We show that Hsp90 possesses two chaperone sites located in the N- and C-terminal fragme nts, respectively. The C-terminal fragment binds to partially folded p roteins in an ATP-independent way potentially regulated by cochaperone s. The N-terminal domain contains a peptide binding site that seems to bind preferentially peptides longer than 10 amino acids. Peptide diss ociation is induced by ATP binding. Furthermore, the antitumor drug ge ldanamycin both inhibits the weak ATPase of Hsp90 and stimulates pepti de release. We propose that the existence of two functionally differen t chaperone sites together with a substrate-selecting set of cochapero nes allows Hsp90 to guide the folding of a subset of target proteins a nd, at the same time, to exhibit general chaperone functions.