Identification of the binding interfaces on CheY for two of its targets, the phosphatase CheZ and the flagellar switch protein FliM

Megan McEvoy, Anat Bren, Michael Eisenbach, Frederick W. Dahlquist

Research output: Contribution to journalArticle

77 Citations (Scopus)

Abstract

CheY is the response regulator protein serving as a phosphorylation-dependent switch in the bacterial chemotaxis signal transduction pathway. CheY has a number of proteins with which it interacts during the course of the signal transduction pathway. In the phosphorylated state, it interacts strongly with the phosphatase CheZ, and also the components of the flagellar motor switch complex, specifically with FliM. Previous work has characterized peptides consisting of small regions of CheZ and FliM which interact specifically with CheY. We have quantitatively measured the binding of these peptides to both unphosphorylated and phosphorylated CheY using fluorescence spectroscopy. There is a significant enhancement of the binding of these peptides to the phosphorylated form of CheY, suggesting that these peptides share much of the binding specificity of the intact targets of the phosphorylated form of CheY. We also have used modem nuclear magnetic resonance methods to characterize the sites of interaction of these peptides on CheY. We have found that the binding sites are overlapping and primarily consist of residues in the C-terminal portion of CheY. Both peptides affect the resonances of residues at the active site, indicating that the peptides may either bind directly at the active site or exert conformational influences that reach to the active site. The binding sites for the CheZ and FliM peptides also overlap with the previously characterized CheA binding interface. These results suggest that interaction with these three proteins of the signal transduction pathway are mutually exclusive. In addition, since these three proteins are sensitive to the phosphorylation state of CheY, it may be that the C-terminal region of CheY is most sensitive for the conformational changes occurring upon phosphorylation.

Original languageEnglish (US)
Pages (from-to)1423-1433
Number of pages11
JournalJournal of Molecular Biology
Volume289
Issue number5
DOIs
StatePublished - Jun 25 1999
Externally publishedYes

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Phosphoric Monoester Hydrolases
Peptides
Proteins
Signal Transduction
Catalytic Domain
Phosphorylation
Binding Sites
Switch Genes
Modems
Fluorescence Spectrometry
Chemotaxis
Magnetic Resonance Spectroscopy

Keywords

  • Chemotaxis
  • CheY
  • CheZ
  • FliM
  • Protein-protein interactions

ASJC Scopus subject areas

  • Virology

Cite this

Identification of the binding interfaces on CheY for two of its targets, the phosphatase CheZ and the flagellar switch protein FliM. / McEvoy, Megan; Bren, Anat; Eisenbach, Michael; Dahlquist, Frederick W.

In: Journal of Molecular Biology, Vol. 289, No. 5, 25.06.1999, p. 1423-1433.

Research output: Contribution to journalArticle

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