In cell extracts of Pseudaminobacter salicylatoxidans strain BN12, an enzymatic activity was detected which converted salicylate in an oxygen-dependent but NAD(P)H-independent reaction to a product with an absorbance maximum at 283 nm. This metabolite was isolated, purified, and identified by mass spectrometry and 1 H and 13 C nuclear magnetic resonance spectroscopy as 2-oxohepta-3,5-dienedioic acid. This metabolite could be formed only by direct ring fission of salicylate by a 1,2-dioxygenase reaction. Cell extracts from P. salicylatoxidans also oxidized 5-aminosalicylate, 3-, 4-, and 5-chlorosalicylate, 3-, 4-, and 5-methylsalicylate, 3-and 5-hydroxysalicylate (gentisate), and 1-hydroxy-2-naphthoate. The dioxygenase was purified and shown to consist of four identical subunits with a molecular weight of about 45,000. The purified enzyme showed higher catalytic constants with gentisate or 1-hydroxy-2-naphthoate than with salicylate. It was therefore concluded that P. salicylatoxidans synthesized a gentisate 1,2-dioxygenase with an extraordinary substrate range, which also allowed the oxidation of salicylate.Naphthalenesulfonic acids are produced on a large scale as industrial detergents, dispersive materials, and intermediates for the production of azo dyes (34). Some pure and mixed bacterial cultures which grow with naphthalene-1-sulfonic acid, naphthalene-2-sulfonic acid (2NS), 2-aminonaphthalene-1-sulfonic acid, 6-aminonaphthalene-2-sulfonic acid (6A2NS), or naphthalenedisulfonic acids as the sole source of carbon and energy have been isolated. The initial reaction in the microbial degradation of (substituted) naphthalenesulfonic acids is catalyzed by desulfonating dioxygenases, which catalyze the formation of (substituted) 1,2-dihydroxynaphthalene(s) and the release of sulfite (4, 5, 26-30, 32, 44). Thus, the degradative pathway for naphthalenesulfonic acids converges at the level of 1,2-dihydroxynaphthalene (1,2-DHN) with the well-known pathway for the degradation of naphthalene (Fig. 1). The reactions which are catalyzed by the "upper" naphthalenesulfonic acid pathway result in the formation of (substituted) salicylate(s). For Sphingomonas xenophaga BN6, it was found that the enzymes participating in the upper naphthalenesulfonic acid pathway convert a wide range of substituted substrates and that therefore a single set of enzymes is sufficient for the conversion of various substituted naphthalenesulfonic acids (19-21, 26, 27, 35). In contrast to the degradation of (substituted) naphthalenesulfonic acids to the corresponding salicylates, the productive metabolism of (substituted) salicylates requires rather different metabolic pathways. Thus, salicylate is usually converted by a salicylate 1-monooxygenase to catechol, 3-hydroxysalicylate (2,3-dihyroxybenzoate) is directly oxidized by an extradiol ring fission reaction to a nonaromatic product, 4-hydroxysalicylate (2,4-dihydroxybenzoate) is converted by a monooxygenase to the ring fission substrate 1,2,4-trihydroxybenzene, and gentisate (5-hydroxysalicylate...