“Molecular basis of the PhoP-PhoQ specific interaction in Salmonella enterica”

E. GARCÍA VÉSCOVI, M. E. CASTELLI, A. CAUERHFF, F. C. SONCINI

Washington D.C., USA

Congreso; 103rd General Meeting, American Society for Microbiology; 2003

American Society for Microbiology

Molecular basis of the PhoP-PhoQ specific interaction in Salmonella enterica García Véscovi, Eleonora, Castelli, María Eugenia, Cauerhff, Ana, and Soncini, Fernando C. IBR-CONICET, School of Biochemistry, Rosario National University, Rosario, and Fundación Instituto Leloir (CONICET-UBA), Buenos Aires, ARGENTINA.   The growing information that proceeds from the analysis of the entire genomic sequences of different bacteria shows that a single cell can harbor up to 40 distinct His-Asp phosphotransfer two-component system pairs.  Though there is a variety of modular arrays in this regulatory family of proteins, in essence, they consist of a sensor protein, typically a histidine kinase (HK), that is responsible for the detection of a specific environmental signal and, through a His-Asp phosphotransfer mechanism, defines the activity state of its associated response regulator (RR).  This RR ultimately controls the generation of an adequate adaptive response, most frequently by modulating gene expression.  In light of the complex network of homologous systems that populate a single bacterial cell, one critical and yet not well-elucidated aspect of the mechanism is the basis for the specificity of the recognition between the HK and its cognate RR.  In the pathogenic bacteria Salmonella enterica serovar Typhimurium the PhoP/PhoQ two-component system governs the adaptation to Mg2+ limited-media.  The detection of this cation allows Salmonella to discern extracellular versus intracellular environments and to control the expression of factors that determine the invasion and spreading in the host.  We recently showed that PhoQ is a bi-functional sensor, demonstrating that the conformational change triggered by Mg2+ promotes a switch in the activity of the sensor from a kinase-dominant to a phosphatase-dominant state.  In this work, we addressed the characterization of the interaction between the two component partners to determine the molecular basis for the fidelity in their recognition.  For this purpose a series of truncated proteins derived from the cytoplasmic region of PhoQ were designed.  By using different in vivo (interference assay using the liberated sensor domains) and in vitro (affinity retention chromatography and surface plasmon resonance) approaches we demonstrated that the PhoQ histidine-phosphotransfer domain is responsible for the specific interaction with PhoP. We also showed that this region undergoes dimerization.