Purification and biochemical characterization of two isolated laccase isoforms from Agaricus bisporus CU13 and their potency in dye decolorization

Abstract

Agaricus bisporus CU13 laccase was purified using ammonium sulfate precipitation (40–80%), Sephadex G100, and DEAE Sephadex A50 anion exchange column chromatography, respectively. Two laccase isoenzymes (Lacc1 & Lacc2) with purification folds of 1.40 and 5.81 respectively, were obtained from DEAE Sephadex A50 column. Optimal temperature and pH were recorded at 55 °C and pH 5.0 for both laccase isoenzymes using ABTS as substrate. Lacc1 was more thermostable than Lacc2 with residual activity of 95, 80 and 6%, while Lacc2 only retained 72, 25 and 0.4% of its activity after incubation for 90 min. at 50, 60 and 70 °C, respectively. Lacc2 retained about 93 and 86% of the initial activity at pH 9.0 and 7.0, whereas Lacc1 was stable at pH 7.0 and 5.0 followed by pH 9.0 and retained about 87, 76, and 36% of its activity respectively, after 4 h of incubation. Lacc1 was activated by 40% in the presence of Cu2+ (10 mM). Km and Vmax values found to be 0.394 and 0.158 μM, and 0.1351 and 0.4755 μmol min−1 for Lacc1 and Lacc2, respectively. The efficiency of both isoenzymes to decolorize Acid blue dye, make the enzyme seems to be a prospective for further biotechnological applications.