Identification of a cis-acting sequence within the Pm promoter of the TOL plasmid which confers XylS-mediated responsiveness to substituted benzoates.

The DNA sequences within the Pm promoter/operator region of the meta operon of the TOL plasmid of Pseudomonas putida, which confer XylS-mediated responsiveness to substituted benzoates, have been identified through deletion analysis and mutagenesis of the region. Integrity and proper phasing of two homologous tandem sequences 5'-TGCAAPuAAPu-PyGGNTA-3', separated by six base-pairs and overlapping with the -35 region of the Pm promoter, was essential for m-toluate activation of a Pm-lacZ fusion in xylS+ strains. The spacing between equivalent bases in each of the half-sites is 21 base-pairs, i.e. two turns of DNA helix. The activity of Pm varieties containing identical half-sites suggested that the XylS-responsive element is arranged as a direct repeat, the distal sequence being the one which provided the most stringent regulation when duplicated. The role of the repeats as the target for XylS binding to Pm is discussed.