The LuxR receptor: the sites of interaction with quorum-sensing signals and inhibitors

• Published in: Microbiology (Society for General Microbiology) 2005-11, Vol.151 (11), p.3589-3602
• Main Authors: Koch, B, Liljefors, T, Persson, T, Nielsen, J, Kjelleberg, S, Givskov, M
• Format: Article
• Language: English
• Subjects: 4-Butyrolactone - analogs & derivatives; 4-Butyrolactone - metabolism; Aliivibrio fischeri - genetics; Aliivibrio fischeri - metabolism; Amino Acid Substitution; Bacteriology; Binding Sites; Biological and medical sciences; Delisea pulchra; Escherichia coli - genetics; Escherichia coli - growth & development; Escherichia coli - metabolism; Fundamental and applied biological sciences. Psychology; Furans - metabolism; Furans - pharmacology; Gene Expression Regulation, Bacterial; Genetics; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; Microbiology; Models, Molecular; Pathogenicity, virulence, toxins, bacteriocins, pyrogens, host-bacteria relations, miscellaneous strains; Repressor Proteins - antagonists & inhibitors; Repressor Proteins - chemistry; Repressor Proteins - genetics; Repressor Proteins - metabolism; Signal Transduction; Trans-Activators - antagonists & inhibitors; Trans-Activators - chemistry; Trans-Activators - genetics; Trans-Activators - metabolism; Vibrio fischeri
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/mic.0.27954-0

1 Center for Biomedical Microbiology, BioCentrum-DTU, Building 301, Technical University of Denmark, DK-2800 Lyngby, Denmark 2 Department of Medicinal Chemistry, The Danish University of Pharmaceutical Sciences, Universitetsparken 2, DK-2100 Copenhagen Ø, Denmark 3 Department of Natural Sciences, The Royal Veterinary and Agricultural University, Thorvaldsensvej 40, DK-1871 Frederiksberg C, Denmark 4 School of Biotechnology and Biomolecular Science and Centre for Marine Biofouling and Bio-innovation, Biological Science Building, University of New South Wales, Randwick, Sydney, NSW 2052, Australia Correspondence Michael Givskov immg{at}pop.dtu.dk The function of LuxR homologues as quorum sensors is mediated by the binding of N -acyl- L -homoserine lactone (AHL) signal molecules to the N-terminal receptor site of the proteins. In this study, site-directed mutagenesis was carried out of the amino acid residues comprising the receptor site of LuxR from Vibrio fischeri , and the ability of the L42A, L42S, Y62F, W66F, D79N, W94D, V109D, V109T and M135A LuxR mutant proteins to activate green fluorescent protein expression from a P luxI promoter was measured. X-ray crystallographic studies of the LuxR homologue TraR indicated that residues Y53 and W57 form hydrogen bonds to the 1-carbonyl group and the ring carbonyl group, respectively, of the cognate AHL signal. Based on the activity and signal specificity of the LuxR mutant proteins, and on molecular modelling, a model is suggested in which Y62 (corresponding to Y53 in TraR) forms a hydrogen bond with the ring carbonyl group rather than the 1-carbonyl group, while W66 (corresponding to W57 in TraR) forms a hydrogen bond to the 1-carbonyl group. This flips the position of the acyl side chain in the LuxR/signal molecule complex compared to the TraR/signal molecule complex. Halogenated furanones from the marine alga Delisea pulchra and the synthetic signal analogue N -(sulfanylacetyl)- L -homoserine lactone can block quorum sensing. The LuxR mutant proteins were insensitive to inhibition by N -(propylsulfanylacetyl)- L -homoserine lactone. In contrast, the mutations had only a minor effect on the sensitivity of the proteins to halogenated furanones, and the data strongly suggest that these compounds do not compete in a ‘classic’ way with N -3-oxohexanoyl- L -homoserine lactone for the binding site. Based on modelling and experimental data it is suggested that these compounds bind in a non-agonist fashion. Abbreviati