[Cloning and expression of Pseudomonas putida gene controlling the catechol-2,3-oxygenase activity in Escherichia coli cells]

Abstract

The genes nahC and nahD from Pseudomonas putida naphthalene degradation plasmid pBS286 were cloned on the vector pUC19 in Escherichia coli cells. The catechol-2,3-oxygenase activity observed in E. coli cells containing recombinant plasmid pBS955 demands the participation of 32 kD polypeptide which is apparently the product of the nahC gene. Second polypeptide of molecular weight 34.5 kD is synthesized in pBS955 containing E. coli minicells and perhaps it is a nahD gene product. The data obtained indicate that 1,2-dihydroxynaphthalene dioxygenase possesses a relaxed substrate specificity, at least, when cloned on a multicopy vector. The cloned DNA insert of pBS955 is not likely to contain its own promoter, so that expression of nahC and nahD genes from the nah1 operon is controlled by the vector lac promoter. The direction of transcription and localization of both polypeptides on the pBS955 map are determined.