Chemotaxis kinase CheA is activated by three neighbouring chemoreceptor dimers as effectively as by receptor clusters.
Chemoreceptors are central to bacterial chemotaxis. These transmembrane homodimers form trimers of dimers. Trimers form clusters of a few to thousands of receptors. A crucial receptor function is 100-fold activation, in signalling complexes, of sensory histidine kinase CheA. Significant activation has been shown to require more than one receptor dimer but the number required for full activation was unknown. We investigated this issue using Nanodiscs, soluble, nanoscale (∼10 nm diameter) plugs of lipid bilayer, to limit the number of neighbouring receptors contributing to activation. Utilizing size-exclusion chromatography, we separated primary preparations of receptor-containing Nanodiscs, otherwise heterogeneous for number and orientation of inserted receptors, into fractions enriched for specific numbers of dimers per disc. Fractionated, clarified Nanodiscs carrying approximately five dimers per disc were as effective in activating kinase as native membrane vesicles containing many neighbouring dimers. At five independently inserted dimers per disc, every disc would have at least three dimers oriented in parallel and thus able act together as they would in native membrane. We conclude full kinase activation involves interaction of CheA with groups of three receptor dimers, presumably as a trimer of dimers, and that more extensive interactions among receptors are not necessary for full kinase activation.
Department of Biochemistry, 117 Schweitzer Hall, University of Missouri-Columbia, Columbia, MO 65211, USA.
Mingshan
M
Li
Cezar M
CM
Khursigara
Sriram
S
Subramaniam
Gerald L
GL
Hazelbauer
D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D005591:Chemical Fractionation; D002633:Chemotaxis; D004789:Enzyme Activation; D004926:Escherichia coli / Q000166:cytology* / Q000201:enzymology*; D029968:Escherichia coli Proteins; D000071677:Histidine Kinase; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D000072236:Methyl-Accepting Chemotaxis Proteins; D049329:Nanostructures / Q000648:ultrastructure; D055503:Protein Multimerization
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; D000071677:Histidine Kinase; C000607608:cheA protein, E coli
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