Previous work has shown that the white rot fungus Coriolopsis rigida degraded wheat straw lignin and both the aliphatic and aromatic fractions of crude oil from contaminated soils. To better understand these processes, we studied the enzymatic composition of the ligninolytic system of this fungus. Since laccase was the sole ligninolytic enzyme found, we paid attention to the oxidative capabilities of this enzyme that would allow its participation in the mentioned degradative processes. We purified two laccase isoenzymes to electrophoretic homogeneity from copper-induced cultures. Both enzymes are monomeric proteins, with the same molecular mass (66 kDa), isoelectric point (3.9), N-linked carbohydrate content (9%), pH optima of 3.0 on 2,6-dimethoxyphenol (DMP) and 2.5 on 2,2-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), absorption spectrum, and N-terminal amino acid sequence. They oxidized 4-anisidine and numerous phenolic compounds, including methoxyphenols, hydroquinones, and lignin-derived aldehydes and acids. Phenol red, an unusual substrate of laccase due to its high redox potential, was also oxidized. The highest enzyme affinity and efficiency were obtained with ABTS and, among phenolic compounds, with 2,6-dimethoxyhydroquinone (DBQH 2 ). The presence of ABTS in the laccase reaction expanded the substrate range of C. rigida laccases to nonphenolic compounds and that of MBQH 2 extended the reactions catalyzed by these enzymes to the production of H 2 O 2 , the oxidation of Mn 2؉ , the reduction of Fe 3؉ , and the generation of hydroxyl radicals. These results confirm the participation of laccase in the production of oxygen free radicals, suggesting novel uses of this enzyme in degradative processes.Lignin is an aromatic heteropolymer of phenyl-propanoid units which confers structural rigidity to woody plant tissues and protects them from microbial attack (25). To depolymerize and mineralize lignin, white rot fungi have developed a nonspecific oxidative system including several extracellular oxidoreductases, low-molecular-weight metabolites, and activated oxygen species (47). The ability of white rot fungi to degrade a wide number of organopollutants is in part due to the action of this nonspecific system (42). Extracellular enzymes involved in the degradation of lignin and xenobiotics by white rot fungi include several kinds of laccases (34, 49), peroxidases (8, 33), and oxidases producing H 2 O 2 (20,32,50).The enzymatic composition of the ligninolytic system depends on the fungal species, with laccase being the common component (24,43). For this reason, a wide number of studies have focused on demonstrating the participation of laccase in significant ligninolytic events which were first attributed to other enzymes of the ligninolytic system. These events include the oxidation of nonphenolic lignin units, which comprise ca. 80% of the polymer, the generation of the H 2 O 2 required for both peroxidase activities and hydroxyl radical ( ⅐ OH) formation, and the production of Mn 3ϩ from the Mn 2ϩ pr...
