Production and characterization of thermostable laccase from the mushroom, Ganoderma lucidum, using submerged fermentation

Abstract

Laccase production by the white rot fungus, Ganoderma lucidum, was investigated using submerged fermentation, and the laccase was purified and characterized. The effects of variations in the media nutrient composition or culture conditions and characteristics of purified laccase were provided for large scale production or industrial uses of the laccase. Low concentrations of glucose and yeast extract stimulated G. lucidum laccase production. The addition of lignocellulosic substrates to the media stimulated laccase production: pomelo peel induced the highest laccase activity of 11842.13 U/L and banana peel reduced laccase production, compared to control media containing wheat bran. Native PAGE and SDS PAGE demonstrated the presence of a single isoform with a molecular weight of 68 kDa. The optimum pH and temperature were 3.0 and 60°C, respectively. G. lucidum laccase is thermostable, with a residual activity of 46% after 80 min at 60°C. The Michaelis-Menten constant (Km) and catalytic constant (Kcat) values for G. lucidum laccase using ABTS (2,2'-azino-di-[3-ethyl-benzothiazolin- sulphonate]) as a substrate were 0.114 mM and 74.63 S-1, respectively, with a specificity constant (Kcat / Km) of 654.65 S-1 mM-1. The thermostable properties of the laccase produced by fermentation of G. lucidum in submerged culture had potential for industrial and biotechnology applications.