Purification and characterization of an endocellulase from the thermophilic fungus Chaetomium thermophilum CT2

Abstract

Chaetomium thermophilum CT2 produced endocellulases at 50 °C, when grown on 2% microcrystalline cellulose, 1% soluble starch, and 0.4% yeast extract medium. A major endocellulase component was purified to homogeneity by fractional ammonium sulphate precipitation, ion-exchange chromatography on DEAE-Sepharose, Phenyl-Sepharose hydrophobic interaction chromatography and gel filtration on Sephacryl S-100. The molecular weight of the enzyme was estimated to be 67.8 kDa and the enzyme was found to be a glycoprotein containing 18.9% carbohydrate. The K m of the purified enzyme for carboxymethyl cellulose, sodium salt (CMC), was 4.6 mg ml −1. The enzyme displayed highest activity towards CMC and significantly lower activities towards phosphoric acid swollen cellulose and filter paper. The activity was enhanced in the presence of Na +, K + and Ca 2+ but inhibited by Hg 2+, Zn 2+, Ag +, Mn 2+, Ba 2+, Fe 2+, Cu 2+, Mg 2+ and NH 4 +. Optimum activity was at 60 °C and pH 4.0. The enzyme was stable over 60 min incubation at 60 °C and half-life at 70, 80 and 90 °C was approximately 45, 24 and 7 min, respectively.