CheW binding interactions with CheA and Tar. Importance for chemotaxis signaling in Escherichia coli.
The initial signaling events underlying the chemotactic response of Escherichia coli to aspartic acid occur within a ternary complex that includes Tar (an aspartate receptor), CheA (a protein kinase), and CheW. Because CheW can bind to CheA and to Tar, it is thought to serve as an adapter protein in this complex. The functional importance of CheW binding interactions, however, has not been investigated. To better define the role of CheW and its binding interactions, we performed biochemical characterization of six mutant variants of CheW. We examined the ability of the purified mutant CheW proteins to bind to CheA and Tar, to promote formation of active ternary complexes, and to support chemotaxis in vivo. Our results indicate that mutations which eliminate CheW binding to Tar (V36M) or to CheA (G57D) result in a complete inability to form active ternary complexes in vitro and render the CheW protein incapable of mediating chemotaxis in vivo. The in vivo signaling pathway can, however, tolerate moderate changes in CheW-Tar and CheW-CheA affinities observed with several of the mutants (G133E, G41D, and 154ocr). One mutant (R62H) provided surprising results that may indicate a role for CheW in addition to binding CheA/receptors and promoting ternary complex formation.
Boukhvalova
Marina S
MS
Department of Cell Biology and Molecular Genetics, University of Maryland, College Park, Maryland 20742, USA.
Dahlquist
Frederick W
FW
Stewart
Richard C
RC
D016880:Anisotropy; D001426:Bacterial Proteins / Q000378:metabolism*; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D002628:Chemoreceptor Cells; D002633:Chemotaxis; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D019793:Fluorescein / Q000494:pharmacology; D005454:Fluorescence Polarization; D000071677:Histidine Kinase; D008565:Membrane Proteins / Q000378:metabolism*; D000072236:Methyl-Accepting Chemotaxis Proteins; D016296:Mutagenesis; D009154:Mutation; D020125:Mutation, Missense; D009841:Oligonucleotides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000378:metabolism*; D015398:Signal Transduction; D020124:Thermotoga maritima / Q000378:metabolism
D001426:Bacterial Proteins; C489103:CheW protein, E coli; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; D009841:Oligonucleotides; D011956:Receptors, Cell Surface; C057331:Tar protein, E coli; C051236:CheW protein, Bacteria; D000071677:Histidine Kinase; C000607608:cheA protein, E coli; D019793:Fluorescein