title: Bacterial sensor kinase TodS interacts with agonistic and antagonistic signals.
abstract: The TodS/TodT two-component system controls expression of the toluene dioxygenase (TOD) pathway for the metabolism of toluene in Pseudomonas putida DOT-T1E. TodS is a sensor kinase that ultimately controls tod gene expression through its cognate response regulator, TodT. We used isothermal titration calorimetry to study the binding of different compounds to TodS and related these findings to their capacity to induce gene expression in vivo. Agonistic compounds bound to TodS and induced gene expression in vivo. Toluene was a powerful agonist, but ortho-substitutions of toluene reduced or abolished in vivo responses, although TodS recognized o-xylene with high affinity. These compounds were called antagonists. We show that agonists and antagonists compete for binding to TodS both in vitro and in vivo. The failure of antagonists to induce gene expression in vivo correlated with their inability to stimulate TodS autophosphorylation in vitro. We propose intramolecular TodS signal transmission, not molecular recognition of compounds by TodS, to be the phenomenon that determines whether a given compound will lead to activation of expression of the tod genes. Molecular modeling identified residues F46, I74, F79, and I114 as being potentially involved in the binding of effector molecules. Alanine substitution mutants of these residues reduced affinities (2- to 345-fold) for both agonistic and antagonistic compounds. Our data indicate that determining the inhibitory activity of antagonists is a potentially fruitful alternative to design specific two-component system inhibitors for the development of new drugs to inhibit processes regulated by two-component systems.
authors: Department of Environmental Protection, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, C/Profesor Albareda, 1, 18008 Granada, Spain.
authors: Andreas
authors: A
authors: Busch
authors: Jesús
authors: J
authors: Lacal
authors: Ariadna
authors: A
authors: Martos
authors: Juan L
authors: JL
authors: Ramos
authors: Tino
authors: T
authors: Krell
mesh_terms: D001426:Bacterial Proteins / Q000819:agonists / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D005786:Gene Expression Regulation; D006838:Hydrocarbons / Q000378:metabolism; D008024:Ligands; D008958:Models, Molecular; D010766:Phosphorylation; D011485:Protein Binding; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D016962:Sinorhizobium meliloti / Q000378:metabolism; D013816:Thermodynamics; D017186:Titrimetry; D014050:Toluene / Q000494:pharmacology; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D001426:Bacterial Proteins; D006838:Hydrocarbons; D008024:Ligands; D047428:Protein Kinase Inhibitors; D011994:Recombinant Proteins; D014050:Toluene; D011494:Protein Kinases
references: Annu Rev Biochem. 2000;69:183-215
references: 10966457
references: J Mol Biol. 2000 Aug 11;301(2):415-31
references: 10926518
references: Mol Biol Evol. 2000 Dec;17(12):1956-70
references: 11110912
references: Nucleic Acids Res. 2001 Jan 1;29(1):55-7
references: 11125048
references: Microbes Infect. 2001 Apr;3(5):417-24
references: 11369279
references: J Biol Chem. 2001 Aug 17;276(33):30827-33
references: 11404360
references: Proteins. 2001;Suppl 5:39-46
references: 11835480
references: Bioorg Med Chem. 2002 Apr;10(4):855-67
references: 11836091
references: Chem Biol. 2002 Nov;9(11):1209-17
references: 12445771
references: J Bacteriol. 2002 Dec;184(24):7062-7
references: 12446657
references: Nat Struct Biol. 2002 Dec;9(12):912-7
references: 12434151
references: J Biol Chem. 2003 May 16;278(20):18434-9
references: 12639952
references: Biochemistry. 2003 May 20;42(19):5917-24
references: 12741850
references: J Bacteriol. 2003 Aug;185(16):4755-63
references: 12896994
references: J Bacteriol. 2003 Aug;185(16):4956-72
references: 12897016
references: Mol Microbiol. 2003 Oct;50(1):219-30
references: 14507376
references: J Biol Chem. 2003 Oct 3;278(40):39189-96
references: 12867417
references: FEMS Microbiol Lett. 2004 Feb 16;231(2):277-81
references: 14987775
references: Curr Med Chem. 2004 Mar;11(6):765-73
references: 15032730
references: J Biol Chem. 1989 Sep 5;264(25):14940-6
references: 2670929
references: J Mol Biol. 1992 May 5;225(1):93-105
references: 1583696
references: Cell. 1996 Jan 12;84(1):165-74
references: 8548821
references: Appl Environ Microbiol. 1997 Feb;63(2):652-60
references: 9023943
references: Proc Natl Acad Sci U S A. 1997 Feb 18;94(4):1453-8
references: 9037074
references: J Bacteriol. 1998 Mar;180(5):1063-71
references: 9495743
references: Gene. 1999 May 17;232(1):69-76
references: 10333523
references: J Bacteriol. 1999 Sep;181(17):5309-16
references: 10464202
references: Trends Microbiol. 2005 Feb;13(2):52-6
references: 15680762
references: J Biol Chem. 2005 May 27;280(21):20887-93
references: 15767250
references: Cell. 2005 Aug 12;122(3):461-72
references: 16096064
references: Nucleic Acids Res. 2006 Jan 1;34(Database issue):D247-51
references: 16381856
references: J Mol Biol. 2006 Mar 10;356(5):1193-206
references: 16406409
references: Microb Ecol. 2006 Feb;51(2):166-76
references: 16435167
references: Proc Natl Acad Sci U S A. 2006 May 23;103(21):8191-6
references: 16702539
references: J Bacteriol. 2006 Jun;188(12):4169-82
references: 16740923
references: Nat Rev Microbiol. 2006 Sep;4(9):705-9
references: 16894339
references: Microbiol Mol Biol Rev. 2006 Dec;70(4):910-38
references: 17158704
references: J Biol Chem. 2007 Jun 1;282(22):16308-16
references: 17416591
references: J Biol Chem. 2000 Dec 8;275(49):38900-4
references: 10978341