Purification of the regulatory protein AlgR1 and its binding in the far upstream region of the algD promoter in Pseudomonas aeruginosa.

AUTOR(ES)

Kato, J

RESUMO

A regulatory protein AlgR1, previously suggested to be a member of a two-component sensory transduction system because of its homology to OmpR and NtrC and its ability to allow activation of the algD promoter under conditions of high osmolarity, has been hyperproduced in Escherichia coli after deletion of the upstream region including part of the Shine-Dalgarno sequence of the algR1 gene and its subsequent cloning under the tac promoter. The AlgR1 protein is purified as a monomer, and the sequence of the nine N-terminal amino acids of the monomer matches with that predicted from the DNA sequence of the algR1 gene. The purified AlgR1 protein binds to two separate DNA fragments of the algD upstream region. DNase protection experiments identify these two DNA segments as 14-mer sequences centered at -382 and -458 regions, which contain a common CCGT-TCGTC sequence in them. While the presence of at least one AlgR1 binding site is important for the activation of the algD promoter, the presence of both of the binding sites in the upstream region leads to a higher level of activation.

Documentos Relacionados

• Role of the far-upstream sites of the algD promoter and the algR and rpoN genes in environmental modulation of mucoidy in Pseudomonas aeruginosa.
• AlgR, a response regulator controlling mucoidy in Pseudomonas aeruginosa, binds to the FUS sites of the algD promoter located unusually far upstream from the mRNA start site.
• Identification of the algZ gene upstream of the response regulator algR and its participation in control of alginate production in Pseudomonas aeruginosa.
• Transcriptional analysis of the Pseudomonas aeruginosa genes algR, algB, and algD reveals a hierarchy of alginate gene expression which is modulated by algT.
• AlgR3, a protein resembling eukaryotic histone H1, regulates alginate synthesis in Pseudomonas aeruginosa.