Urea-dependent signal transduction by the virulence regulator UreR

Inessa Gendlina, Delia M. Gutman, Venetta Thomas, Carleen M. Collins

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

Identification of the environmental triggers involved in the expression of virulence genes is a fundamental objective in studies of bacterial pathogens. For uropathogens, urea, found in the urinary tract at concentrations of up to 500 mM, functions as an environmental signal. Urea freely diffuses into the bacterium Providencia stuartii and activates UreR, a member of the AraC family of transcriptional activators. Active UreR promotes transcription of virulence-associated urease genes and alerts the organisms of its immediate milieu. Thus, the UreR.urea complex has a dual role, acting as both a transcriptional activator as well as an environmental sensor. Here, we describe the molecular events associated with activation of gene expression by urea-bound UreR. The Kd of the urea·UreR binding reaction was measured as 0.2 mM by fluorescence quenching assays, and the shape of the binding curve indicated a single specific urea-binding site on UreR. Histidine residues are critical for urea binding in urease, and therefore to identify the urea-binding site in UreR, five mutant UreR forms were generated with histidine to alanine substitutions. Two of the mutants (UreRc) exhibited a constitutive phenotype by both activating transcription and binding to DNA with an increased affinity in the absence of urea. The UreRc bound urea with an affinity similar to that of wild-type UreR. We concluded, therefore, that the mutations resulting in constitutive activity were not involved in the UreR.urea interaction. UreR was activated, then, either by binding urea or by histidine to alanine substitutions at one of two positions. Circular dichroism indicated little change in the structure of UreR when activated, and size-exclusion chromatography demonstrated that both rUreR and rUreRc were dimers in both the presence and absence of urea. Thus, the structural changes associated with activation are subtle.

Original languageEnglish (US)
Pages (from-to)37349-37358
Number of pages10
JournalJournal of Biological Chemistry
Volume277
Issue number40
DOIs
StatePublished - Oct 4 2002
Externally publishedYes

Fingerprint

Signal transduction
Virulence
Urea
Signal Transduction
Histidine
Urease
Transcription
Alanine
Substitution reactions
Genes
Chemical activation
Binding Sites
Providencia
Gene Expression
Size exclusion chromatography
Pathogens
Circular Dichroism
Urinary Tract
Gene expression
Dimers

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Urea-dependent signal transduction by the virulence regulator UreR. / Gendlina, Inessa; Gutman, Delia M.; Thomas, Venetta; Collins, Carleen M.

In: Journal of Biological Chemistry, Vol. 277, No. 40, 04.10.2002, p. 37349-37358.

Research output: Contribution to journalArticle

Gendlina, Inessa ; Gutman, Delia M. ; Thomas, Venetta ; Collins, Carleen M. / Urea-dependent signal transduction by the virulence regulator UreR. In: Journal of Biological Chemistry. 2002 ; Vol. 277, No. 40. pp. 37349-37358.
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