PRODUCTION OF SUBSTITUTED NAPHTHALENE DIHYDRODIOLS BY ENGINEERED ESCHERICHIA-COLI CONTAINING THE CLONED NAPHTHALENE 1,2-DIOXYGENASE GENE FROM PSEUDOMONAS-FLUORESCENS N3

Naphthalene dioxygenase, a key enzyme in the dihydroxylation of naphth alene, is encoded by the plasmid pN3, responsible for naphthalene meta bolism in Pseudomonas fluorescens N3. The naphthalene dioxygenase, inc luding all the sequences for its expression and the regulatory region, has been localized on the 4.3-kb HindIII-ClaI fragment and on the 3.5 -kb HindIII fragment of the plasmid pN3, by Southern analysis using as probes nahA and nahR genes, the homologous genes of the plasmid NAH7 from Pseudomonas putida G7. We cloned in Escherichia coli JM109 the di oxygenase gene and its regulatory region and developed an efficient ba cterial system inducible by salicylic acid, able to produce dihydrodio ls. E. coli containing recombinant plasmids carrying the dioxygenase g ene were analysed for their potential as a biocatalytic tool to produc e dihydrodiols from different naphthalenes with the substituent on the aromatic ring at the alpha or beta position. The dihydrodiols, identi fied by HPLC (high-performance liquid chromatography) and H-1-NMR (nuc lear magnetic resonance) were produced with yields ranging from 50 to 94%. The degree of bioconversion efficiency depends on the nature and the position of the substituent and indicates the broad substrate spec ificity of this dioxygenase and its potential for the production of a wide variety of fine chemicals.