Carbon catabolite repression of phenol degradation in Pseudomonas putida is mediated by the inhibition of the activator protein PhlR

Abstract

Enzymes involved in (methyl)phenol degradation of Pseudomonas putida H are encoded by the catabolic operon (phlA-L) on plasmid pPGH1. Transcription of this operon by the sigma54 (RpoN)-containing RNA polymerase is positively controlled by the gene product of the divergently transcribed phlR in response to the availability of the respective substrate. Additionally, phenol degradation is subject to carbon catabolite repression induced by organic acids (e.g., succinate, lactate, and acetate) or carbohydrates (e.g., glucose and gluconate). Analysis of lacZ fusion to the catabolic promoter and quantified primer extension experiments indicate that carbon catabolite repression also occurs at the transcriptional level of the catabolic operon. In this study, it is furthermore shown that carbon catabolite repression is a negative control. Titration of the postulated negative controlling factor was exclusively observed when extra copies of functional phlR gene were present in the cell. We therefore conclude that PhlR is the target and that carbon catabolite repression of phenol degradation occurs by interfering with the activating function of PhlR.