Activation of the transcriptional regulator XylR of Pseudomonas putida by release of repression between functional domains.
In the presence of toluene, xylenes and other structural analogues, the regulatory protein XylR, of the family of transcriptional regulators which act in concert with the sigma 54 factor, activate the promoter Pu of the TOL (toluene degradation) plasmid pWWO of Pseudomonas putida. Amino acid changes Val-219-Asp and Ala-220-Pro, introducing a proline kink at the hinge region between the N-terminal A domain and the central portion of XylR, resulted in a semi-constitutive phenotype which mimicked the activating effect of aromatic inducers. This phenotype was further exacerbated by inserting extra amino acid residues within the same inter-domain region. A truncated XylR protein devoid of the signal-receiving, amino-terminal portion of the protein stimulated the cognate promoter Pu at high levels independently of inducer addition, both in Escherichia coli and in Pseudomonas putida. Replacement of the amino-terminal domain by a heterologous peptide derived from the MS2 virus polymerase resulted in a hybrid protein still able to bind DNA to the same extent in vivo as XylR, but unable to stimulate transcription. These data indicate that a key event in the activation of XylR by toluene/xylenes is the release of the repression caused by the A domain of the protein on surfaces located at the central domain of the regulator.
Fernández
S
S
Centro de Investigaciones Biológicas CSIC, Madrid, Spain.
de Lorenzo
V
V
Pérez-Martín
J
J
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics*; D004268:DNA-Binding Proteins; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005809:Genes, Regulator; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010641:Phenotype; D011401:Promoter Regions, Genetic; D016958:Pseudomonas putida / Q000235:genetics*; D015398:Signal Transduction; D014157:Transcription Factors; D014158:Transcription, Genetic; D014992:Xylenes / Q000378:metabolism
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D014157:Transcription Factors; C074322:XylR protein, Pseudomonas; D014992:Xylenes; C031285:3-xylene