Targeting of the chemotaxis methylesterase/deamidase CheB to the polar receptor-kinase cluster in an Escherichia coli cell.
Chemotactic adaptation to persisting stimulation involves reversible methylation of the chemoreceptors that form complexes with the histidine kinase CheA at a cell pole. The methyltransferase CheR targets to the C-terminal NWETF sequence of the chemoreceptor. In contrast, localization of the methylesterase CheB is largely unknown, although regulation of its activity via phosphorylation is central to adaptation. In this study, green fluorescent protein was fused to full-length CheB or its various parts: the N-terminal regulatory domain (N), the C-terminal catalytic domain (C) and the linker (L). The full-length and NL fusions and, to a lesser extent, the LC fusion localized to a pole. Deletion of the P2 domain from CheA abolished polar localization of the full-length and NL fusions, but did not affect that of the LC fusion. Pull-down assays demonstrated that the NL fragment, but not the LC fragment, binds to the P2 fragment of CheA. These results indicate that binding of the NL domain to the P2 domain targets CheB to the polar signalling complex. The LC fusion, like the chemoreceptor, partially localized in the absence of CheA, suggesting that the LC domain may interact with its substrate sites, either as part of the protein or as a proteolytic fragment.
Banno
Satomi
S
Division of Biological Science, Graduate School of Science, Nagoya University, Chikusa-ku, Nagoya 464-8602, Japan.
Shiomi
Daisuke
D
Homma
Michio
M
Kawagishi
Ikuro
I
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016764:Cell Polarity; D002633:Chemotaxis; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D049452:Green Fluorescent Proteins; D000071677:Histidine Kinase; D008164:Luminescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D000072236:Methyl-Accepting Chemotaxis Proteins; D008958:Models, Molecular; D005810:Multigene Family; D016297:Mutagenesis, Site-Directed; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013347:Subcellular Fractions
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008164:Luminescent Proteins; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; D011993:Recombinant Fusion Proteins; C069656:CheB protein, Bacteria; D049452:Green Fluorescent Proteins; D011494:Protein Kinases; D000071677:Histidine Kinase; C000607608:cheA protein, E coli; D002265:Carboxylic Ester Hydrolases; C041144:chemotactic protein methylesterase