Structure of the constitutively active double mutant CheY D13K Y106W alone and in complex with a FliM peptide

Collin M. Dyer, Michael L. Quillin, Andres Campos, Justine Lu, Megan M. McEvoy, Andrew C. Hausrath, Edwin M. Westbrook, Philip Matsumura, Brian W. Matthews, Frederick W. Dahlquist

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35 Scopus citations

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 1.5 Å resolution with an R-factor of 19.6%. The asymmetric unit contains four CheY** molecules, two with FliM 16 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.

Original languageEnglish (US)
Pages (from-to)1325-1335
Number of pages11
JournalJournal of Molecular Biology
Volume342
Issue number4
DOIs
StatePublished - Sep 24 2004

Keywords

  • CheY
  • FliM
  • activating mutation
  • chemotaxis
  • protein-protein interactions

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

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    Dyer, C. M., Quillin, M. L., Campos, A., Lu, J., McEvoy, M. M., Hausrath, A. C., Westbrook, E. M., Matsumura, P., Matthews, B. W., & Dahlquist, F. W. (2004). Structure of the constitutively active double mutant CheY D13K Y106W alone and in complex with a FliM peptide. Journal of Molecular Biology, 342(4), 1325-1335. https://doi.org/10.1016/j.jmb.2004.07.084