Principal Investigator Tania Baker
Protein Processing During Disassembly and Degradation
Several groups studying Clp/Hsp100 proteins demonstrated that these enzymes have the capacity to unfold their substrate proteins completely. My group completed two studies of this type with ClpXP. We found that the rate of unfolding of the stable model substrate was accelerated by 7 orders of magnitude by ClpX. Furthermore, experiments with proteins of various intrinsic stabilities strongly supported the conclusion that unfolding occurs via an active mechanism, rather than by passive trapping of the transiently unfolded state. Unfolding is essential for ClpXP-mediated protein degradation, as the entry pore to the ClpP protease chamber is ~10 Å, too small to allow passage of anything other than an unfolded polypeptide.
With the knowledge of the ClpX mechanism based on degradation studies, we returned to studying the mechanism used by ClpX to destabilize the Mu transposase–DNA complex. Several lines of experiments reveal that protein unfolding by ClpX is involved in disassembly, but that only some of the subunits in the complex are unfolded. Thus, we conclude that the ability of ClpX to unfold proteins is sufficient to explain its role in both protein destabilization and ATP-dependent proteolysis.