Study on Purification and Some Properties of A Highly Glucose-tolerant Extracellular β-Glucosidase from Aspergillus niger

Two kind of extracellular β-glucosidases were purified to homogeneity from an Aspergillus niger by ammonium sulfate precipitation,Phenyl Sepharose CL-4B hydrophobic interaction chromatography,DEAE-Sepharose ion exchange chromatography and Sephacryl S-200 HR gel layer chromatography.One of them was a highly glucose-tolerant novel β-glucosidase with a Ki of 41.01 mmol/L,and the final purification factor 56.7 times and 22.66 % yield were obtained.The enzyme single subunit mole-cular weight was about 114.6 ku which was identified by SDS-PAGE.The Km and vmax values of the enzyme were 0.904 mmol/L and 1.08 μmol/min,respectively,using p-nitrophenyl-β-D-glucopyranoside(pNPG) as a substrate.The optimum reaction temperature and pH value for β-glucosidase were 60 ℃ and pH value 4.0,respectively.The enzyme was stable in the pH value range of 3.0-7.0 and up to 60 ℃.The enzyme was greatly inhibited by Ag+.Other metal ions tested and EDTA hand no effects on the activity of β-glucosidase.Different organic solvent had different effects on the activity of β-glucosidase,methanol and 1-butanol enhanced the activity,whereas acetone and acetonitrile inhibited the activity obviously.