title: Structure of the constitutively active double mutant CheYD13K Y106W alone and in complex with a FliM peptide.
abstract: CheY is a member of the response regulator protein superfamily that controls the chemotactic swimming response of motile bacteria. The CheY double mutant D13K Y106W (CheY**) is resistant to phosphorylation, yet is a highly effective mimic of phosphorylated CheY in vivo and in vitro. The conformational attributes of this protein that enable it to signal in a phosphorylation-independent manner are unknown. We have solved the crystal structure of selenomethionine-substituted CheY** in the presence of its target, a peptide (FliM16) derived from the flagellar motor switch, FliM, to 1.5A resolution with an R-factor of 19.6%. The asymmetric unit contains four CheY** molecules, two with FliM16 bound, and two without. The two CheY** molecules in the asymmetric unit that are bound to FliM16 adopt a conformation similar to BeF3- -activated wild-type CheY, and also bind FliM16 in a nearly identical manner. The CheY** molecules that do not bind FliM16 are found in a conformation similar to unphosphorylated wild-type CheY, suggesting that the active phenotype of this mutant is enabled by a facile interconversion between the active and inactive conformations. Finally, we propose a ligand-binding model for CheY and CheY**, in which Ile95 changes conformation in a Tyr/Trp106-dependent manner to accommodate FliM.
authors.affiliation: Institute of Molecular Biology, University of Oregon, Eugene, OR 97403, USA.
authors.forename: Collin M
authors.initials: CM
authors.lastname: Dyer
authors.forename: Michael L
authors.initials: ML
authors.lastname: Quillin
authors.forename: Andres
authors.initials: A
authors.lastname: Campos
authors.forename: Justine
authors.initials: J
authors.lastname: Lu
authors.forename: Megan M
authors.initials: MM
authors.lastname: McEvoy
authors.forename: Andrew C
authors.initials: AC
authors.lastname: Hausrath
authors.forename: Edwin M
authors.initials: EM
authors.lastname: Westbrook
authors.forename: Philip
authors.initials: P
authors.lastname: Matsumura
authors.forename: Brian W
authors.initials: BW
authors.lastname: Matthews
authors.forename: Frederick W
authors.initials: FW
authors.lastname: Dahlquist
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000072236:Methyl-Accepting Chemotaxis Proteins; D008958:Models, Molecular; D009154:Mutation; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D015398:Signal Transduction; D014443:Tyrosine / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; C066821:FliM protein, Bacteria; D014443:Tyrosine