title: The structures of T87I phosphono-CheY and T87I/Y106W phosphono-CheY help to explain their binding affinities to the FliM and CheZ peptides.
abstract: CheY is a response regulator in bacterial chemotaxis. Escherichia coli CheY mutants T87I and T87I/Y106W CheY are phosphorylatable on Asp57 but unable to generate clockwise rotation of the flagella. To understand this phenotype in terms of structure, stable analogs of the two CheY-P mutants were synthesized: T87I phosphono-CheY and T87I phosphono-CheY. Dissociation constants for peptides derived from flagellar motor protein FliM and phosphatase CheZ were determined for phosphono-CheY and the two mutants. The peptides bind phosphono-CheY almost as strongly as CheY-P; however, they do not bind T87I phosphono-CheY or T87I/Y106W phosphono-CheY, implying that the mutant proteins cannot bind FliM or CheZ tightly in vivo. The structures of T87I phosphono-CheY and T87I/Y106W phosphono-CheY were solved to resolutions of 1.8 and 2.4A, respectively. The increased bulk of I87 forces the side-chain of Y106 or W106, into a more solvent-accessible conformation, which occludes the peptide-binding site.
authors.affiliation: Department of Chemistry and Biochemistry, University of North Carolina Wilmington, 601 S. College Road, Wilmington, NC 28403, USA.
authors.forename: Kenneth
authors.initials: K
authors.lastname: McAdams
authors.forename: Eric S
authors.initials: ES
authors.lastname: Casper
authors.forename: R
authors.initials: R
authors.lastname: Matthew Haas
authors.forename: Bernard D
authors.initials: BD
authors.lastname: Santarsiero
authors.forename: Aimee L
authors.initials: AL
authors.lastname: Eggler
authors.forename: Andrew
authors.initials: A
authors.lastname: Mesecar
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Halkides
mesh_terms: D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites / Q000502:physiology; D002633:Chemotaxis / Q000502:physiology; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000072236:Methyl-Accepting Chemotaxis Proteins; D020125:Mutation, Missense; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology; D020836:Protein Structure, Quaternary / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; D010455:Peptides; C000607606:cheY protein, E coli; C066821:FliM protein, Bacteria
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