TRANSPOSON AND SPONTANEOUS DELETION MUTANTS OF PLASMID-BORNE GENES ENCODING POLYCYCLIC AROMATIC HYDROCARBON DEGRADATION BY A STRAIN OF PSEUDOMONAS-FLUORESCENS

Pseudomonas fluorescens strain LP6a, isolated from petroleum condensat e-contaminated soil, utilizes the polycyclic aromatic hydrocarbons (PA Hs) naphthalene, phenanthrene, anthracene and 2-methylnaphthalene as s ole carbon and energy sources. The isolate also co-metabolically trans forms a suite of PAHs and heterocycles including fluorene, biphenyl, a cenaphthene, 1-methylnaphthalene, indole, benzothiophene, dibenzothiop hene and dibenzofuran, producing a variety of oxidized metabolites. A 63 kb plasmid (pLP6a) carries genes encoding enzymes necessary for the PAH-degrading phenotype of P. fluorescens LP6a. This plasmid hybridiz es to the classical naphthalene degradative plasmids NAH7 and pWW60, b ut has different restriction endonuclease patterns. In contrast, plasm id pLP6a failed to hybridize to plasmids isolated from several phenant hrene-utilizing strains which cannot utilize naphthalene. Plasmid pLP6 a exhibits reproducible spontaneous deletions of a 38 kb region contai ning the degradative genes. Two gene clusters corresponding to the arc hetypal naphthalene degradation upper and lower pathway operons, separ ated by a cryptic region of 18 kb, were defined by transposon mutagene sis. Gas chromatographic-mass spectrometric analysis of metabolites ac cumulated by selected transposon mutants indicates that the degradativ e enzymes encoded by genes on pLP6a have a broad substrate specificity permitting the oxidation of a suite of polycyclic aromatic and hetero cyclic substrates.