title: Structural Basis of an N-Degron Adaptor with More Stringent Specificity.
abstract: The N-end rule dictates that a protein's N-terminal residue determines its half-life. In bacteria, the ClpS adaptor mediates N-end-rule degradation, by recognizing proteins bearing specific N-terminal residues and delivering them to the ClpAP AAA+ protease. Unlike most bacterial clades, many α-proteobacteria encode two ClpS paralogs, ClpS1 and ClpS2. Here, we demonstrate that both ClpS1 and ClpS2 from A. tumefaciens deliver N-end-rule substrates to ClpA, but ClpS2 has more stringent binding specificity, recognizing only a subset of the canonical bacterial N-end-rule residues. The basis of this enhanced specificity is addressed by crystal structures of ClpS2, with and without ligand, and structure-guided mutagenesis, revealing protein conformational changes and remodeling in the substrate-binding pocket. We find that ClpS1 and ClpS2 are differentially expressed during growth in A. tumefaciens and conclude that the use of multiple ClpS paralogs allows fine-tuning of N-end-rule degradation at the level of substrate recognition.
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Benjamin J
authors.initials: BJ
authors.lastname: Stein
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Robert A
authors.initials: RA
authors.lastname: Grant
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Robert T
authors.initials: RT
authors.lastname: Sauer
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Howard Hughes Medical Institute, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. Electronic address: tabaker@mit.edu.
authors.forename: Tania A
authors.initials: TA
authors.lastname: Baker
mesh_terms: D016960:Agrobacterium tumefaciens / Q000737:chemistry / Q000254:growth & development* / Q000378:metabolism; D000596:Amino Acids / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018360:Crystallography, X-Ray; D015964:Gene Expression Regulation, Bacterial; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010447:Peptide Hydrolases / Q000378:metabolism; D011487:Protein Conformation; D058767:Protein Unfolding; D013379:Substrate Specificity
chemical_list: D000596:Amino Acids; D001426:Bacterial Proteins; D002352:Carrier Proteins; D010447:Peptide Hydrolases