Suppression of a mutation in OmpR at the putative phosphorylation center by a mutant EnvZ protein in Escherichia coli.
Phosphorylation of OmpR, a transcription activator for ompF and ompC expression, is essential for its function and has been shown to be mediated in vitro by EnvZ, a transmembrane sensory receptor protein. On the basis of the three-dimensional structure of CheY which has an extensive sequence similarity with OmpR, three aspartic residues, D11, D12, and D55, of OmpR are considered to form a triacidic pocket serving as the phosphorylation center. When these aspartic acid residues were replaced with asparagine (D11N) or glutamine (D12Q and D55Q), ompF and ompC expression was almost completely blocked. Two pseudorevertants of the D11N mutation were isolated: one of them is a mutation in EnvZ (G240E), and the other is a mutation in OmpR (S48F). The envZ mutation (G240E) by itself was found to confer a phenotype very similar to that of the well known envZ11 mutation (T247R), suggesting that EnvZ (G240E) is an elevated kinase for OmpR. Consistent with this notion, EnvZ (T247R) was also able to suppress the D11N mutation in OmpR. An in vitro phosphorylation study showed that while the wild-type OmpR was phosphorylated by EnvZ, the D11N OmpR was not. These results suggest that the D11N mutation alters OmpR conformation in such a way that OmpR is very poorly phosphorylated by EnvZ. On the basis of the in vivo and in vitro analysis, the mechanisms by which the G240E mutation in EnvZ and the S48F mutation in OmpR suppress the D11N mutation in OmpR are discussed.
Brissette
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RE
Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Rutgers, Piscataway 08854.
Tsung
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KL
Inouye
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D001425:Bacterial Outer Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D029968:Escherichia coli Proteins; D019811:Hydroxylamine; D006898:Hydroxylamines / Q000494:pharmacology; D008958:Models, Molecular; D009097:Multienzyme Complexes; D016296:Mutagenesis; D010766:Phosphorylation; D010957:Plasmids; D011487:Protein Conformation; D013489:Suppression, Genetic; D014158:Transcription, Genetic
D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; D006898:Hydroxylamines; D009097:Multienzyme Complexes; D019811:Hydroxylamine; C057843:envZ protein, E coli