Mycosystema. 2008, 27(2): 297-308.
Three kinds of laccases from Bjerkandera adusta WZFF.W-Y11 were purified to electrophoretic homogeneity by the combination of acetone fractionation, DEAE-Cellulose ion exchange, and Sephadex G-100 chromatography. They were identified by the color development of zymogram stained with guaiacol after polyacrylamide gel electrophoresis (PAGE), and named LacA, LacB and LacC, respectively. The enzymes were purified 29.2-folds, 5.1-folds and 18.5-folds respectively from crude samples with a total recovery yield of 65.5%. The laccases were enzymologically characterized and the results showed that the molecular weight of LacA, LacB and LacC were 68.7kDa, 80.2kDa, and 77.2kDa, respectively, as determined by SDS-polyacrylamide gel electrophoresis (SDS-PAGE). The Km of both LacA and LacC for oxidizing guaiacol were higher than that for oxidizing ABTS. Their optimum pH and temperature were 3.0-5.5 and 45℃-70℃, respectively. At 65℃, LacC showed a better stability than LacA. The activity of LacA was relatively stable at pH range of 5.0-9.0, and LacC at pH range of 3.5-6.0. Various ions had different effects on laccase activity, which was enhanced by Al3+, but strongly inhibited by Hg2+, Fe3+ and Cl-. Cu2+ had no effect on LacA activity but inhibited LacC activity.
