title: Mutational analysis of the N-terminal domain of UreR, the positive transcriptional regulator of urease gene expression.
abstract: The Escherichia coli plasmid-encoded urease, a virulence factor in human and animal infections of the urinary and gastroduodenal tracts, is induced when the substrate urea is present in the growth medium. Urea-dependent urease expression is mediated at the transcriptional level by the AraC-like activator UreR. Previous work has shown that a peptide representing the N-terminal 194 amino-acid residues of UreR binds urea at a single site, full-length UreR forms an oligomer, and the oligomerization motif is thought to reside in the N-terminal portion of the molecule. The C-terminal domain of UreR contains two helix-turn-helix motifs presumed to be necessary for DNA binding. In this study, we exploited mutational analyses at the N-terminal domain of UreR to determine if this domain dimerizes similar to other AraC family members. UreR mutants were analyzed for the ability to activate transcription of lacZ from an ureDp-lacZ transcriptional fusion. A construct encoding the N-terminal 194 amino acids of UreR, eluted as an oligomer by gel filtration and had a dominant negative phenotype over the wild-type ureR allele. We hypothesize that this dominant negative phenotype results from the formation of inactive heterodimers between wild-type and truncated UreR. Dominant negative analysis and cross-linking assays demonstrated that E. coli UreR is active as a dimer and dimerization occurs within the first 180 residues.
authors.affiliation: Department of Microbiology, University of Washington, Seattle, WA 98195, United States. consuelo.parra@gmail.com
authors.forename: Maria C
authors.initials: MC
authors.lastname: Parra
authors.forename: Carleen M
authors.initials: CM
authors.lastname: Collins
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004252:DNA Mutational Analysis; D019281:Dimerization; D004926:Escherichia coli / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014510:Urease / Q000235:genetics* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D015534:Trans-Activators; C119767:UreR protein, bacteria; D014510:Urease