Characterization of the Bacterial Sensor Protein PhoQ

The PhoP/PhoQ two-component regulatory system governs several virulence properties in the Gram-negative bacterium Salmonella typhimurium. The PhoQ protein is a Mg2+ and Ca2+ sensor that modulates transcription of PhoP-regulated genes in response to the extracellular concentrations of these divalent cations. We have purified a 146-amino acid polypeptide corresponding to the periplasmic (i.e. sensing) domain of the PhoQ protein. Mg2+ altered the tryptophan intrinsic fluorescence of this polypeptide whereas Ba2+, which is unable to modulate transcription of PhoP-regulated genes, did not. Mg2+ was more effective than Ca2+ at repressing transcription of PhoP-activated genes in vivo. However, maximal repression was achieved when both cations were present. An avirulent mutant harboring a single amino acid substitution in the sensing domain of PhoQ exhibited lower affinity for Ca2+ but similar affinity for Mg2+. Cumulatively, these experiments demonstrate that Mg2+ can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg2+ and Ca2+ in the PhoQ protein.

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