Identification of the repressor subdomain within the signal reception module of the prokaryotic enhancer-binding protein XylR of Pseudomonas putida

In the presence of m-xylene, the protein XylR encoded by the TOL plasmid of Pseudomonas putida, activates the sigma(54)-dependent promoter Pu. Early activation stages involve the release of the intramolecular repression caused by the signal reception N-terminal (A domain) of XylR on the central module of the protein. A genetic approach has been followed to locate the specific segment within A domain of XylR that is directly responsible for its down-regulation in the absence of inducer, as compared to that involved in effector (m-xylene) binding. For this, a reporter Escherichia coli strain carrying a monocopy transcriptional fusion of Pu to lacZ was transformed with a collection of plasmids encoding equivalent truncated varieties of XylR, consisting of nested and internal deletions throughout the entire A domain. Examination of the resulting phenotypes allowed the assignment of the A domain region near the central activation domain, as the portion of the protein responsible for the specific repression of XylR activity in the absence of m-xylene.