Purification and characterization of an extracellular β-glucosidase II with high hydrolysis and transglucosylation activities from Aspergillus niger

An extracellular beta-glucosidase II (beta-Glu II) has been purified to homogeneity by column chromatography from Aspergillus niger CCRC 31494. Its molecular mass was estimated to be 360 kDa by gel filtration and 120 kDa by SDS-PAGE. The enzyme has a pI of 4.0 and has optimum activity at pH 4.5 and 60 degrees C. The beta-Glu II was completely inhibited by 5.0 mM Fe2+. Methanol (20%, v/v) activated the enzyme activity at 1.8-fold. V-max values of 10.2 and 464 units/mg were found for p-nitrophenyl beta-D-glucoside (K-m = 2.2 mM) and for cellobiose (K-m = 15.4 mM). The enzyme was strongly inhibited by substrates, p-nitrophenyl beta-D-glucopyranoside in excess of 7.5 mM and cellobiose in excess of 50 mM, and was competitively inhibited by glucose with a K-i of 5.7 mM. Transglucosylation products of cellotriose, methyl beta-glucoside and ethyl beta-glucoside, were obtained under neutral conditions and in the presence of methanol and ethanol, respectively.