title: Remodeling of a delivery complex allows ClpS-mediated degradation of N-degron substrates.
abstract: The ClpS adaptor collaborates with the AAA+ ClpAP protease to recognize and degrade N-degron substrates. ClpS binds the substrate N-degron and assembles into a high-affinity ClpS-substrate-ClpA complex, but how the N-degron is transferred from ClpS to the axial pore of the AAA+ ClpA unfoldase to initiate degradation is not known. Here we demonstrate that the unstructured N-terminal extension (NTE) of ClpS enters the ClpA processing pore in the active ternary complex. We establish that ClpS promotes delivery only in cis, as demonstrated by mixing ClpS variants with distinct substrate specificity and either active or inactive NTE truncations. Importantly, we find that ClpA engagement of the ClpS NTE is crucial for ClpS-mediated substrate delivery by using ClpS variants carrying "blocking" elements that prevent the NTE from entering the pore. These results support models in which enzymatic activity of ClpA actively remodels ClpS to promote substrate transfer, and highlight how ATPase/motor activities of AAA+ proteases can be critical for substrate selection as well as protein degradation.
authors.affiliation: Department of Biology and.
authors.forename: Izarys
authors.initials: I
authors.lastname: Rivera-Rivera
authors.affiliation: Department of Biology and.
authors.forename: Giselle
authors.initials: G
authors.lastname: Román-Hernández
authors.affiliation: Department of Biology and.
authors.forename: Robert T
authors.initials: RT
authors.lastname: Sauer
authors.affiliation: Department of Biology and Howard Hughes Medical Institute, Massachusetts Institute of Technology, Cambridge, MA 02139 tabaker@mit.edu.
authors.forename: Tania A
authors.initials: TA
authors.lastname: Baker
mesh_terms: D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D049071:Endopeptidase Clp / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D050505:Mutant Proteins / Q000378:metabolism; D059748:Proteolysis; D013379:Substrate Specificity
chemical_list: D002352:Carrier Proteins; C454820:ClpS protein, E coli; D029968:Escherichia coli Proteins; D050505:Mutant Proteins; C483568:ClpA protease, E coli; D049071:Endopeptidase Clp
keywords: AAA+ ATPase adaptor; AAA+ unfoldase/translocase; N-degron substrate selection; adaptor remodeling