Laccase (EC 1.10.3.2) from the culture filtrate of a strain of white rot basidiomycete Pleurotus ostreatus was purified using DEAE-Toyopearl 650M and butyI-Toyopearl 650M column chromatographies and Superdex 75 HR 10/30 fast protein liquid chromatography. Molecular weight of the purified laccase was about 55,000, and the isoelectric point was 3.0. The optimum pH for enzyme activity was 6.5, and the optimum temperature was 50~This enzyme contained 7.4% sugar and two copper atoms per molecule. The substrate specificity was similar to those of other fungal laccases. Comparison of the N-terminal amino acid sequence of the P, ostreatus laccase with those from Pleurotus ostreatus Florida, Coriolus hirsutus, Phlebia radiata, basidiomycete PM1 (CECT 2971), Trametes villosa, Pycnoporus cinnabarinus, Ceriporiopsis subvermispora, and Agaricus bisporus showed 95, 65, 60, 55, 55, 55, 50, and 35% similarity, respectively, in the first 20 residues. No similarity in this region was detected with laccases from Neurospora crassa, Aspergillus nidulans, and Cryptococcus neoformans.Key Words--basidiomycete; laccase; Pleurotus ostreatus.Microorganisms with degradation systems of complex lignocellulosics are potentially applicable for detoxification of various organic compounds that cause environmental pollution. Among the ligninolytic enzymes of basidiomycetes, such as laccase, lignin peroxidase, and manganese peroxidase, we have studied laccases, which are found in major white-rot fungi capable of degrading lignin. Extracellular laccases have been purified and characterized from such fungi as Trametes versicolor (Cheung and Marshall, 1969;F~ihraeus and Reinhammar, 1967;Mosbach, 1963;Rogalski et al., 1990), Ganoderma lucidum (Kumari and Sirsi, 1972), Neurospora crassa (Froehner and Eriksson, 1974), Agaricus bisporus (Wood, 1980), Aspergillus nidulans (Kurtz and Champe, 1982), Schizophyllum commune (de Vries et al., 1986), Lentinus edodes (Kofujita et al., 1991 ), Panus tigrinus (Maltseva et al., 1991), Armillaria mellea (Rehman and Thurston, 1992), basidiomycete PM 1 (CECT 2971) (Coil et al., 1993), Cryptococcus neoformans (Williamson, 1994), Trametes villosa (Yaver et al., 1996), and Pycnoporus cinnabarinus (Eggert et al., 1996). Though these laccases have similar molecular weights and similar substrate specificities, they exhibit considerably different amino acid sequences.In our diagnostic studies on phenoloxidase (laccase) activities of various edible basidiomycetes, 24 strains of *Corresponding author, Present address: Department of Biotechnology, Faculty of Engineering, Tottori University, 4-101 Koyama, Tottori 680-8552, Japan; E-maih okarnoto(~bio. tottori u.ac.jp.Pleurotus ostreatus (hiratake in Japanese) all showed remarkably high activities, in agreement with the report on European mushrooms (Bollag and Leonowicz, 1984). A monokaryotic P. ostreatus line, K16-2, which we isolated from the protoplasts of a wild dikaryotic strain in Japan, was found to be one of the most active strains with laccases. In our previous study, we a...