Purification and Characterization of β-Glucosidase Produced by Trichoderma Citrinoviride Cultivated on Microalga Chlorella Vulgaris

Microalga Chlorella vulgaris has a wide application as a source of biomass fuel, natural food coloring agent and dietary supplement. Its cell wall consists of multiple layers including cellulose and in order to utilize microalgae, the pre-treatment is necessary to breakdown rigid cell walls. beta-glucosidase (EC 3.2.1.21) is a crucial enzyme to hydrolyze cellulose efficiently. In this study, beta-glucosidase produced by Trichoderma citrinoviride cultivated on microalgae C. vulgaris was purified to homogeneity with a recovery of 8.5% and specific activity of 168.7 U/mg. The purified enzyme was obtained as a single band with the molecular mass of 110 kDa on SDS-PAGE. The optimum pH and temperature for enzyme activity and stability were 4.0 and 50 degrees C, and 8.0 and 30 degrees C, respectively. Metal ions (Mg2+, and Zn2+) activated the enzyme activity, whereas SDS inhibited moderately. K-m , V-max, K-cat, and K-i values of beta-glucosidase were 0.96 mM, 300.42 mu mol min(-1) mg(-1), 2.73 min(-) (1) and 2.83 mM using p-nitrophenyl beta-D-glucoside as a substrate, whereas they were 4.30 mM, 24.34 mu mol min(-1) mg(-1), 0.022 min(-1) and 0.53 mM using cellobiose, respectively.