Purification and characterization of an extracellular b-glucosidase with high transglucosylation activity and stability from Aspergillus niger No. 5.1.
An extracellular beta-glucosidase was extracted from the culture filtrate of Aspergillus niger No. 5.1 and purified to homogeneity by using ammonium sulfate precipitation, Chitopearl-DEAE chromatography, and Sephadex G-100 chromatography. The specific activity of the enzyme was enriched 6.33-fold, with a recovery of 11.67%. The enzyme was a monomer and the molecular mass was 67.5 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis and 66.5 kDa by gel-filtration chromatography. The enzyme had optimum activity at pH 6.0 and 60 degrees C and was stable over the pH range of 3.0-9.0. It showed specificity of hydrolysis for p-nitrophenyl-beta-D-glucoside and cellobiose. The Km and Vmax values of the enzyme for cellobiose and salicin were 5.34 mM, 2.57 micromol/(mL.s), and 3.09 mM, 1.34 micromol/(mL.s), respectively. Both amino acid composition and N-terminal amino acid sequence of the enzyme were determined, which provides useful information for cloning of this enzyme.
Xie
Yu
Y
Technical Institute of Physics and Chemistry, Chinese Academy of Sciences, No. 3 Datun Road, Beijing 100101, P.R. China.
Gao
Yunhua
Y
Chen
Zhifeng
Z
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001234:Aspergillus niger / Q000201:enzymology*; D004795:Enzyme Stability; D006031:Glycosylation; D007700:Kinetics; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D001617:beta-Glucosidase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism
D000596:Amino Acids; D001617:beta-Glucosidase