Targeting and activation of the AAA+ chaperone ClpB by cooperating Hsp70 during protein disaggregation

The hexameric AAA+ chaperone ClpB rescues proteins from an aggregated state, an activity that is essential for thermotolerance development. Cooperation with an Hsp70 chaperone system is essential for ClpB disaggregation activity. Hsp70 acts first during the initial stages of the disaggregation reaction, followed by substrate threading through the central pore of the ClpB ring. The mechanism of chaperone cooperation and the requirement for Hsp70 at the start of the disaggregation process is not understood. Here, we demonstrate that Hsp70 acts as a targeting factor, allowing for the recruitment of ClpB to protein aggregates. Furthermore, we show that ClpB exist in two functional states, a basal one with low unfolding power and an activated state with high unfolding power. The activity state is controlled by the ClpB-specific M-domain, which is contacting the first AAA domain, thereby controlling ATPase activity. Mutants that arrest the protein in each state exhibit a specific loss of disaggregation activity. The M-domain is also directly mediating the cooperation with Hsp70. We therefore suggest that Hsp70 interaction triggers conformational changes within the M-domain, leading to the activation of ClpB at the surface of protein aggregates.