Kanchan, K, Linder, J, Winkler, K, Hantke, K, Schultz, A and Schultz, JE (2010) Transmembrane signaling in chimeras of the Escherichia coli aspartate and serine chemotaxis receptors and bacterial class III adenylyl cyclases. J. Biol. Chem. 285:2090-9
The Escherichia coli chemoreceptors for serine (Tsr) and aspartate (Tar) and several bacterial class III adenylyl cyclases (ACs) share a common molecular architecture; that is, a membrane anchor that is linked via a cytoplasmic HAMP domain to a C-terminal signal output unit. Functionality of both proteins requires homodimerization. The chemotaxis receptors are well characterized, whereas the typical hexahelical membrane anchor (6TM) of class III ACs, suggested to operate as a channel or transporter, has no known function beyond a membrane anchor. We joined the intramolecular networks of Tsr or Tar and two bacterial ACs, Rv3645 from Mycobacterium tuberculosis and CyaG from Arthrospira platensis, across their signal transmission sites, connecting the chemotaxis receptors via different HAMP domains to the catalytic AC domains. AC activity in the chimeras was inhibited by micromolar concentrations of l-serine or l-aspartate in vitro and in vivo. Single point mutations known to abolish ligand binding in Tar (R69E or T154I) or Tsr (R69E or T156K) abrogated AC regulation. Co-expression of mutant pairs, which functionally complement each other, restored regulation in vitro and in vivo. Taken together, these studies demonstrate chemotaxis receptor-mediated regulation of chimeric bacterial ACs and connect chemical sensing and AC regulation.
Adenylate Cyclase/antagonists & inhibitors; Adenylate Cyclase/chemistry; Adenylate Cyclase/metabolism; Amino Acid Sequence; Aspartic Acid/pharmacology; Bacterial Proteins/chemistry; Bacterial Proteins/metabolism; Cell Membrane/metabolism; Chemoreceptor Cells/chemistry; Chemoreceptor Cells/metabolism; Cyanobacteria/enzymology; Escherichia coli/cytology; Escherichia coli/enzymology; Escherichia coli/metabolism; Escherichia coli Proteins/chemistry; Escherichia coli Proteins/metabolism; Membrane Proteins/chemistry; Membrane Proteins/metabolism; Molecular Sequence Data; Mycobacterium tuberculosis/enzymology; Protein Structure, Tertiary; Receptors, Cell Surface; Recombinant Fusion Proteins/chemistry; Recombinant Fusion Proteins/metabolism; Sequence Alignment; Serine/pharmacology; Signal Transduction
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