Intramolecular second-site revertants to the phosphorylation site mutation in OmpR, a kinase-dependent transcriptional activator in Escherichia coli.
OmpR is a transcriptional activator for the ompF and ompC genes of Escherichia coli. Its phosphorylation is mediated by a transmembrane sensory-receptor protein, EnvZ, and is essential for transcriptional activation. In a previous study, when the aspartic acid residue at position 55, the putative phosphorylation site, was replaced with glutamine (D55Q), ompF and ompC expression were completely lost. In this study two pseudorevertants of the D55Q mutation were isolated and identified to be the replacement of threonine at position 83 with alanine (T83A) and glycine at position 94 with serine (G94S). The revertant OmpRs no longer responded to EnvZ function when ompF and ompC expression were examined. The purified D55Q-T83A OmpR was unable to be phosphorylated by EnvZ in vitro. The role of EnvZ as an osmosensor for the environmentally regulated expression of OmpF and OmpC has been indicated in previous studies. The isolation of seemingly EnvZ-independent OmpR revertants in this study, however, made it possible to examine the osmolarity-regulated expression of OmpF and OmpC in the absence of effects exerted by EnvZ. We found that the expression of OmpF and OmpC supported by these revertant OmpRs was clearly regulated in accordance with the change in osmolarity of the growth media. These results indicate that another EnvZ-independent mechanism(s) may also contribute to the regulated expression of the ompF and ompC genes.
Brissette
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RE
Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Rutgers, Piscataway 08854.
Tsung
K L
KL
Inouye
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M
D001425:Bacterial Outer Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D016172:DNA Fingerprinting; D004269:DNA, Bacterial / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes; D009154:Mutation; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D010957:Plasmids; D011487:Protein Conformation; D015534:Trans-Activators / Q000235:genetics*
D001425:Bacterial Outer Membrane Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D009097:Multienzyme Complexes; D015534:Trans-Activators; C057843:envZ protein, E coli; D010749:Phosphoprotein Phosphatases