Kinase activity of EnvZ, an osmoregulatory signal transducing protein of Escherichia coli

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

EnvZ is an inner membrane protein present in Escherichia coli that is important for osmosensing and required for porin gene regulation. EnvZ is phosphorylated by intracellular ATP, and EnvZ-P phosphorylates OmpR, which then binds to the porin promoters to regulate their expression. An overexpressed, truncated form of the enzyme, EnvZ115, was used to characterize the kinase reaction in vitro. Using a filter binding assay, we report the first direct measurements of the kinase activity, including the apparent affinity for ATP of 200 μM. The phosphorylation reaction is dependent on MgCl2, and the phosphoenzyme has the expected stability of a phosphohistidine; i.e., it is stable in base and less stable in acid at room temperature. The addition of OmpR and ATP to solutions containing EnvZ resulted in an OmpR-stimulated, EnvZ-dependent ATPase activity that was not vanadate-sensitive. The in vivo kinase activity of EnvZ and two mutants that were deficient in porin expression were studied using an immune complex kinase reaction. Interestingly, a mutation located in the periplasmic domain of EnvZ exhibited kinase activity that was identical to that of the wild- type enzyme, while a mutation located close to the phosphorylation site showed a significant decrease in both kinase and phosphotransferase activities. These data provide support for models of EnvZ consisting of separate sensing and kinase domains.

Original languageEnglish (US)
Pages (from-to)303-311
Number of pages9
JournalArchives of Biochemistry and Biophysics
Volume346
Issue number2
DOIs
StatePublished - Oct 15 1997
Externally publishedYes

Fingerprint

Escherichia coli Proteins
Phosphotransferases
Porins
Phosphorylation
Adenosine Triphosphate
Mutation
Magnesium Chloride
Vanadates
Enzymes
Antigen-Antibody Complex
Gene expression
Escherichia coli
Adenosine Triphosphatases
Assays
Membrane Proteins
Temperature
Acids
Genes

Keywords

  • EnvZ
  • Escherichia coli
  • OmpR
  • Osmoregulation
  • Porins
  • Response regulator
  • Two-component regulatory system

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

Cite this

Kinase activity of EnvZ, an osmoregulatory signal transducing protein of Escherichia coli. / Kenney, Linda.

In: Archives of Biochemistry and Biophysics, Vol. 346, No. 2, 15.10.1997, p. 303-311.

Research output: Contribution to journalArticle

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