Mutational analysis of the Escherichia coli PhoQ sensor kinase: differences with the Salmonella enterica serovar Typhimurium PhoQ protein and in the mechanism of Mg2+ and Ca2+ sensing.
The PhoP-PhoQ two-component system plays a role in Mg2+ homeostasis and/or the virulence properties of a number of bacterial species. A Salmonella enterica serovar Typhimurium PhoQ sensor kinase mutant, in which the threonine at residue 48 in the periplasmic sensor domain is changed to an isoleucine, was shown previously to result in elevated expression of PhoP-activated genes and to affect mouse virulence, epithelial cell invasion, and sensitivity to macrophage killing. We characterized a complete set of proteins having amino acid substitutions at position 48 in the closely related Escherichia coli PhoQ protein. Numerous mutant proteins having amino acid substitutions with side chains of various sizes and characters displayed signaling phenotypes similar to that of the wild-type protein, indicating that interactions mediated by the wild-type threonine side chain are not required for normal protein function. Changes to amino acids with aromatic side chains had little impact on signaling in response to extracellular Mg2+ but resulted in reduced sensitivity to extracellular Ca2+, suggesting that the mechanisms of signal transduction in response to these two divalent cations are different. Surprisingly, the Ile48 protein displayed a defective phenotype rather than the hyperactive phenotype seen with the S. enterica serovar Typhimurium protein. We also describe a mutant PhoQ protein lacking the extracellular sensor domain with a defect in the ability to activate PhoP. The defect does not appear to be due to reduced autokinase activity but rather appears to be due to an effect on the stability of the aspartyl-phosphate bond of phospho-PhoP.
Regelmann
Adam G
AG
Department of Microbiology, College of Physicians and Surgeons, Columbia University, New York, New York 10032, USA.
Lesley
Joseph A
JA
Mott
Christina
C
Stokes
Lissette
L
Waldburger
Carey D
CD
D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D004252:DNA Mutational Analysis; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D008274:Magnesium / Q000378:metabolism; D012486:Salmonella typhimurium / Q000201:enzymology* / Q000235:genetics; D015398:Signal Transduction
D001426:Bacterial Proteins; C079792:PhoQ protein, Bacteria; C061064:PhoP protein, Bacteria; D008274:Magnesium; D002118:Calcium