Characterization of the bacterial sensor protein PhoQ: evidence for distinct binding sites for Mg2+ and Ca2+

GARCÍA VÉSCOVI, E.; AYALA, Y; DI CERA, E; GROISMAN, E.A.

JOURNAL OF BIOLOGICAL CHEMISTRY

American Society for Biochemistry and Molecular Biology

Año: 1997 vol. 272 p. 1440 - 1443

0021-9258

The PhoP/PhoQ two-component regulatory system governs several virulence properties in the Gram-negative bacterium Salmonella typhimurium. The PhoQ protein is aMg21 and Ca21 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. Mg21 altered the tryptophan intrinsic fluorescence of this polypeptide whereas Ba21, which is unable to modulate transcription of PhoP-regulated genes, did not. Mg21 was more effective than Ca21 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222in 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222. 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222222i.e. sensing) domain of the PhoQ protein. Mg21 altered the tryptophan intrinsic fluorescence of this polypeptide whereas Ba21, which is unable to modulate transcription of PhoP-regulated genes, did not. Mg21 was more effective than Ca21 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222in 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222. 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.2222222Salmonella typhimurium. The PhoQ protein is aMg21 and Ca21 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. Mg21 altered the tryptophan intrinsic fluorescence of this polypeptide whereas Ba21, which is unable to modulate transcription of PhoP-regulated genes, did not. Mg21 was more effective than Ca21 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222in 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222. 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222222i.e. sensing) domain of the PhoQ protein. Mg21 altered the tryptophan intrinsic fluorescence of this polypeptide whereas Ba21, which is unable to modulate transcription of PhoP-regulated genes, did not. Mg21 was more effective than Ca21 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222in 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222. 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222222221 and Ca21 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. Mg21 altered the tryptophan intrinsic fluorescence of this polypeptide whereas Ba21, which is unable to modulate transcription of PhoP-regulated genes, did not. Mg21 was more effective than Ca21 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222in 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222. 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222222i.e. sensing) domain of the PhoQ protein. Mg21 altered the tryptophan intrinsic fluorescence of this polypeptide whereas Ba21, which is unable to modulate transcription of PhoP-regulated genes, did not. Mg21 was more effective than Ca21 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222in 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222. 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.22222221 altered the tryptophan intrinsic fluorescence of this polypeptide whereas Ba21, which is unable to modulate transcription of PhoP-regulated genes, did not. Mg21 was more effective than Ca21 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222in 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222. 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.2222221, which is unable to modulate transcription of PhoP-regulated genes, did not. Mg21 was more effective than Ca21 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222in 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222. 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222221 was more effective than Ca21 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222in 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.222. 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 Ca21 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.22221 but similar affinity for Mg21. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.2221. Cumulatively, these experiments demonstrate that Mg21 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.221 can bind to the sensing domain of PhoQ and establish the presence of distinct binding sites for Mg21 and Ca21 in the PhoQ protein.21 and Ca21 in the PhoQ protein.