A closed translocation channel in the substrate-free AAA+ ClpXP protease diminishes rogue degradation

Abstract

ABSTRACTIntracellular proteases must be specific to avoid degrading the wrong proteins. Here, we present cryo-EM structures of E. coli ClpXP, a AAA+ protease, which reveal that the axial channel of ClpX is closed prior to the binding and subsequent translocation of a protein substrate. An open-channel ClpX mutation stimulates degradation of casein, a non-specific substrate, indicating that channel closure contributes to increased degradation specificity. We demonstrate that ClpX activates ClpP cleavage of a degron-free decapeptide by a channel-independent mechanism, in which the peptide substrate appears to pass through a symmetry mismatched gap in the interface between ClpX and ClpP before entering the degradation chamber via the axial portal of ClpP. The peptide products of ClpXP protein degradation are likely to exit the chamber by the reverse route.