Conserved aspartate residues and phosphorylation in signal transduction by the chemotaxis protein CheY.
The CheY protein is phosphorylated by CheA and dephosphorylated by CheZ as part of the chemotactic signal transduction pathway in Escherichia coli. Phosphorylation of CheY has been proposed to occur on an aspartate residue. Each of the eight aspartate residues of CheY was replaced by using site-directed mutagenesis. Substitutions at Asp-12, Asp-13, or Asp-57 resulted in loss of chemotaxis. Most of the mutant CheY proteins were still phosphorylated by CheA but exhibited modified biochemical properties, including reduced ability to accept phosphate from CheA, altered phosphate group stability, and/or resistance to CheZ-mediated dephosphorylation. The properties of CheY proteins bearing a substitution at position 57 were most aberrant, consistent with the hypothesis that Asp-57 is the normal site of acyl phosphate formation. Evidence for an alternate site of phosphorylation in the Asp-57 mutants is presented. Phosphorylated CheY is believed to cause tumbling behavior. However, a dominant mutant CheY protein that was not phosphorylated in vitro caused tumbling in vivo in the absence of CheA. This phenotype suggests that the role of phosphorylation in the wild-type CheY protein is to stabilize a transient conformational change that can generate tumbling behavior.
Division of Biology, California Institute of Technology, Pasadena 91125.
R B
RB
Bourret
J F
JF
Hess
M I
MI
Simon
D001224:Aspartic Acid; D001426:Bacterial Proteins; D002630:Chemotactic Factors / Q000235:genetics*; D002633:Chemotaxis; D004926:Escherichia coli / Q000235:genetics* / Q000502:physiology; D029968:Escherichia coli Proteins; D000071677:Histidine Kinase; D007700:Kinetics; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D000072236:Methyl-Accepting Chemotaxis Proteins; D008954:Models, Biological; D009154:Mutation; D010766:Phosphorylation; D015398:Signal Transduction
D001426:Bacterial Proteins; D002630:Chemotactic Factors; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; C000607606:cheY protein, E coli; D001224:Aspartic Acid; D000071677:Histidine Kinase; C000607608:cheA protein, E coli