Mating behavior of Pseudomonas mendocina KR1 and cloning and characterization of its p-cresol utilizing genes.

Abstract

The genes encoding the toluene degradative pathway of Pseudomonas mendocina KR1 were found to be chromosomally encoded and could be transferred by conjugation from the chromosome of strain KR1 to the chromosome of a Pseudomonas aeruginosa or Pseudomonas putida recipient. The transferred genes enabled the recipients to metabolize toluene, p-cresol, p-hydroxybenzaldehyde, and p-hydroxybenzoate. The Toluene$\sp+$ recipients were able to transfer the genes encoding toluene metabolism only when sex-factor was provided in trans. The genes encoding the metabolism of toluene to protocatechuate, the ring cleavage substrate, in P. mendocina KR1 were found to be organized into three separately regulated units: one encoding toluene-4-monooxygenase; a second encoding p-cresol methylhydroxylase and p-hydroxybenzaldehyde dehydrogenase; and a third encoding p-hydroxybenzoate hydroxylase. The genes encoding p-cresol methylhydroxylase and p-hydroxybenzaldehyde dehydrogenase were organized as an operon, in which the gene encoding p-hydroxybenzaldehyde dehydrogenase was transcribed first, followed by transcription of the genes encoding p-cresol methylhydroxylase. The operon was regulated by a positively-acting regulator. p-Hydroxybenzaldehyde dehydrogenase activity was detected in Escherichia coli under control of the tac promoter; however, p-cresol methylhydroxylase was detected only in P. aeruginosa and not in E. coli. The gene encoding p-hydroxybenzoate hydroxylase from strain KR1 was also cloned and was regulated independent of the genes encoding p-cresol methylhydroxylase, p-hydroxybenzaldehyde dehydrogenase, or toluene-4-monooxygenase. The toluene metabolizing pathway of strain KR1 could have been assembled through the acquisition of the individual genes encoding the enzymes of the pathway. The genes encoding the pathway are sufficiently linked to permit mobilization of the genes to other Pseudomonas species. The ability to mobilize genes encoding a catabolic pathway and the expression of those genes in different genetic backgrounds permits the dissemination of that pathway within a microbial community and the rapid adaptation of the microbial community for substate metabolism.