BINDING OF NONNATIVE PROTEIN TO HSP25 DURING HEAT-SHOCK CREATES A RESERVOIR OF FOLDING INTERMEDIATES FOR REACTIVATION

Small heat shock proteins (sHsps) are a conserved and ubiquitous prote in family. Their ability to convey thermoresistance suggests their par ticipation in protecting the native conformation of proteins. However, the underlying functional principles of their protective properties a nd their role in concert with other chaperone families remain enigmati c, Here, we analysed the influence of Hsp25 on the inactivation and su bsequent aggregation of a model protein, citrate synthase (CS), under heat shock conditions in vitro. We show that stable binding of several non-native CS molecules to one Hsp25 oligomer leads to an accumulatio n of CS unfolding intermediates, which are protected from irreversible aggregation. Furthermore, a number of different proteins which bind t o Hsp25 can he isolated from heat-shocked extracts of cells. Under per missive folding conditions, CS can be released from Hsp25 and, in coop eration with Hsp70, an ATP-dependent chaperone, the native state can b e restored. Taken together, our findings allow us to integrate sHsps f unctionally in the cellular chaperone system operating under heat shoc k conditions. The task of sHsps in this context is to efficiently trap a large number of unfolding proteins in a folding-competent state and thus create a reservoir of non-native proteins for an extended period of time, allowing refolding after restoration of physiological condit ions in cooperation with other chaperones.