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BIOLOGICAL SCIENCES / MICROBIOLOGY
Bacterial sensor kinase TodS interacts with agonistic and antagonistic signals
*Department of Environmental Protection, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, C/Profesor Albareda, 1, 18008 Granada, Spain; and Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, C/Ramiro de Maetzu, 9, 28040 Madrid, Spain
Edited by Thomas J. Silhavy, Princeton University, Princeton, NJ, and approved June 26, 2007 (received for review February 23, 2007)
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.
histidine kinases | isothermal titration calorimetry | Pseudomonas | two-component systems | aromatic hydrocarbons
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
This article contains supporting information online at www.pnas.org/cgi/content/full/0701547104/DC1.
To whom correspondence should be addressed. E-mail: jlramos{at}eez.csic.es
© 2007 by The National Academy of Sciences of the USA
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