Protein Remodeling by ClpB AAA+ ATPase Independent of the DnaK/Hsp70 Chaperone System

Abstract

AAA+ ATPases forcibly remodel diverse macromolecular structures. ClpB, a member of the AAA+ family of ATPases and a Clp/Hsp100 chaperone, is a hexameric protein containing two AAA+ domains and an axial channel spanning the length of the hexamer. ClpB is required for cell survival after exposure to extreme thermal stress and functions by dissolving and renaturing aggregated proteins. ClpB acts in protein disaggregation in conjunction with the DnaK chaperone and its two co-chaperones, DnaJ and GrpE, both in vivo and in vitro. Remarkably, when given mixtures of ATP and ATP gamma S (a poorly hydrolysable ATP analog), ClpB is activated to perform protein remodeling functions independent of the DnaK chaperone system in vitro. This finding has allowed us to study ClpB activities in isolation, without the complicating effects of DnaK, DnaJ and GrpE. We have compared ClpB to other Clp/Hsp100 proteins, including ClpA and ClpX, in protein activation using the model “small aggregate”, P1 RepA, in protein unfolding using GFP fusion proteins, and in protein renaturation using heat-aggregated proteins. We have found that in some aspects ClpB is like ClpA and ClpX and in others it is somewhat different.