Bacterial Energy Sensor Aer Modulates the Activity of the Chemotaxis Kinase CheA Based on the Redox State of the Flavin Cofactor.
Flagellated bacteria modulate their swimming behavior in response to environmental cues through the CheA/CheY signaling pathway. In addition to responding to external chemicals, bacteria also monitor internal conditions that reflect the availability of oxygen, light, and reducing equivalents, in a process termed "energy taxis." In Escherichia coli, the transmembrane receptor Aer is the primary energy sensor for motility. Genetic and physiological data suggest that Aer monitors the electron transport chain through the redox state of its FAD cofactor. However, direct biochemical data correlating FAD redox chemistry with CheA kinase activity have been lacking. Here, we test this hypothesis via functional reconstitution of Aer into nanodiscs. As purified, Aer contains fully oxidized FAD, which can be chemically reduced to the anionic semiquinone (ASQ). Oxidized Aer activates CheA, whereas ASQ Aer reversibly inhibits CheA. Under these conditions, Aer cannot be further reduced to the hydroquinone, in contrast to the proposed Aer signaling model. Pulse ESR spectroscopy of the ASQ corroborates a potential mechanism for signaling in that the resulting distance between the two flavin-binding PAS (Per-Arnt-Sim) domains implies that they tightly sandwich the signal-transducing HAMP domain in the kinase-off state. Aer appears to follow oligomerization patterns observed for related chemoreceptors, as higher loading of Aer dimers into nanodiscs increases kinase activity. These results provide a new methodological platform to study Aer function along with new mechanistic details into its signal transduction process.
From the Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853 and.|the National Biomedical Center for Advanced ESR Technologies, Cornell University, Ithaca, New York 14853.
Dipanjan
D
Samanta
From the Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853 and.
Joanne
J
Widom
From the Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853 and.|the National Biomedical Center for Advanced ESR Technologies, Cornell University, Ithaca, New York 14853.
Peter P
PP
Borbat
From the Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853 and.|the National Biomedical Center for Advanced ESR Technologies, Cornell University, Ithaca, New York 14853.
Jack H
JH
Freed
From the Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853 and bc69@cornell.edu.
Brian R
BR
Crane
D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000071677:Histidine Kinase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D036341:Intercellular Signaling Peptides and Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D000072417:Protein Domains
C108679:Aer protein, E coli; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D036341:Intercellular Signaling Peptides and Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; D005182:Flavin-Adenine Dinucleotide; D000071677:Histidine Kinase; C000607608:cheA protein, E coli
Nanodiscs; bacterial chemotaxis; bacterial energy taxis; chemotaxis; electron transport system (ETS); flavin adenine dinucleotide (FAD); oxidation-reduction (redox); redox signaling