Identification of the DNA binding sites of PerA, the transcriptional activator of the bfp and per operons in enteropathogenic Escherichia coli.
J Bacteriol
; 185(9): 2835-47, 2003 May.
Article
em En
| MEDLINE
| ID: mdl-12700263
ABSTRACT
The bundle-forming pilus (BFP) is an important virulence factor for enteropathogenic Escherichia coli (EPEC). Genes involved in its biogenesis and regulation are tightly regulated by PerA (BfpT), a member of the AraC/XylS family of transcriptional regulators. The aim of this work was to purify PerA and determine its association with bfpA and perA (bfpT) regulatory regions by electrophoretic mobility shift and DNase I footprinting assays. PerA was purified as a maltose-binding protein (MBP) fusion, which was capable of complementing bfpA expression and which was able to restore the localized adherence phenotype of an EPEC perA mutant strain. Upstream of bfpA and perA, MBP-PerA recognized with similar affinity asymmetric nucleotide sequences in which a 29-bp-long AT-rich consensus motif was identified. These DNA motifs share 66% identity and were previously shown, by deletion analysis, to be involved in the PerA-dependent expression of both genes. Interestingly, in perA, this motif spans the sequence between positions -75 and -47, approximately one helix turn upstream of the -35 promoter sequence, while in bfpA, it spans the sequence between positions -83 and -55, approximately two helix turns upstream from the promoter. An additional PerA binding site was identified at the 5' end of the bfpA structural gene, which was not required for its activation. Experiments with LexA-PerA fusions suggested that PerA acts as a monomer to activate the transcription of both perA and bfpA, in contrast to what has been documented for other members of this family of transcriptional regulators.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Óperon
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Proteínas Repressoras
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Proteínas de Escherichia coli
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Proteínas de Fímbrias
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Escherichia coli
Tipo de estudo:
Diagnostic_studies
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Prognostic_studies
Idioma:
En
Ano de publicação:
2003
Tipo de documento:
Article