KINETIC CHARACTERIZATION OF PURIFIED LACCASE PRODUCED FROM Trametes versicolor IBL-04 IN SOLID STATE BIO-PROCESSING OF CORNCOBS

Muhammad Asgher, Hafiz M. Nasir Iqbal, M. Javaid Asad, M. Javaid Asad

Abstract


A locally isolated white rot fungal strain Trametes versicolor IBL-04 produced high laccase activities in solid state bio-processing of corn cobs. Addition of glucose and yeast extract (C: N ratio; 25:1) enhanced laccase synthesis. Addition of Tween-80 and CuSO4 enhanced laccase production to 1012 U/mL under optimized process conditions. Laccase was further purified to 2.89-fold (specific activity of 840 U/mg) by ammonium sulfate fractional precipitation, dialysis, and Sephadex G-100 gel filtration chromatography. The purified laccase had a relative molecular mass of 63 kDa as detected by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). Best enzyme activity was at pH 5 and 40oC. Using 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) as substrate, the enzyme showed maximum activity (Vmax) of 780 U/mL with a corresponding Michaelis constant (Km) value of 73µM. Among the different activators/inhibitors, Cu2+, Mn2+, and Fe2+ stimulated laccase activity, whereas EDTA and cystein inhibited the enzyme. The higher Vmax and lower Km for T. versicolor IBL-04 laccase as compared to most of the reported laccases suggests its potential for industrial applications.

Keywords


Trametes versicolor IBL-04; Corn cobs; Extracellular laccase; Purification; Kinetic studies

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