Copyright © 1994, American Society for Microbiology pH Dependence of CheA Autophosphorylation in Escherichia coli

Chemotaxis by cells of Escherichia coli and SalmoneUla typhimurium depends upon the ability of chemore-ceptors called transducers to communicate with switch components of flagellar motors to modulate swimming behavior. This communication requires an excitatory pathway composed of the cytoplasmic signal transduction proteins, CheAL, CheAs, CheW, CheY, and CheZ. Of these, the autokinase CheAL is most central. Modifications or mutations that affect the rate at which CheAL autophosphorylates result in profound chemotactic defects. Here we demonstrate that pH can affect CheAL autokinase activity in vitro. This activity exhibits a bell-shaped dependence upon pH within the range 6.5 to 10.0, consistent with the notion that two proton dissociation events affect CheAL autophosphorylation kinetics: one characterized by a pK, of about 8.1 and another exhibiting a pK. of about 8.9. These in vitro results predict a decrease in the rate of CheAL autophosphorylation in response to a reduction in intracellular pH, a decrease that should cause increased counterclockwise flagellar rotation. We observed such a response in vivo for cells containing a partially reconstituted chemotaxis system. Benzoate (10 mM, pH 7.0), a weak acid that when undissociated readily traverses the cytoplasmic membrane, causes a reduction of cytoplasmic pH from 7.6 to 7.3. In response to this reduction, cells expressing CheAL, CheAs, and CheY, but not transducers, exhibited a small but reproducible increase in the fraction of time that they spun their flagellar motors counterclockwise. The added presence o