2,3-Dihydroxybenzoate meta -Cleavage Pathway is Involved in o -Phthalate Utilization in Pseudomonas sp. strain PTH10

Pseudomonas sp. strain PTH10 can utilize o- phthalatewhich is a key intermediate in the bacterial degradation of some polycyclic aromatic hydrocarbons. In this strain, o- phthalateis degraded to 2,3-dihydroxybenzoate and further metabolized via the 2,3-dihydroxybenzoate meta-cleavage pathway. Here, the opa genes which are involved in the o- phthalate catabolismwere identified. Based on the enzymatic activity of the opa gene products, opaAaAbAcAd, opaB, opaC, and opaD were found to code for o- phthalate2,3- dioxygenase, dihydrodiol dehydrogenase, 2,3-dihydroxybenzoate 3,4- dioxygenase, and 3-carboxy-2-hydroxymuconate-6-semialdehyde decarboxylase, respectively. Collectively, these enzymes are thought to catalyze the conversion of o- phthalateto 2-hydroxymuconate-6-semialdehyde. Deletion mutants of the above opa genes indicated that their products were required for the utilization of o- phthalate. Transcriptional analysis showed that the opa genes were organized in the same transcriptional unit. Quantitative analysisof opaAa, opaB, opaC, opaD, opaE, and opaN revealed that, except for opaB and opaC, all other genes were transcriptionally induced during growth on o- phthalate. The constitutive expression of opaB and opaC, and the transcriptional induction of opaD located downstream of opaB, suggest several possible internal promoters are existence in the opa cluster. Together, these results strongly suggest that the opa genes are involved in a novel o- phthalate catabolicpathway in strain PTH10.