Purification and characterization of a beta-glucosidase from Aspergillus niger.
The high-molar mass form of beta-glucosidase from Aspergillus niger strain NIAB280 was purified to homogeneity with a 46-fold increase in purification by a combination of ammonium sulfate precipitation, hydrophobic interaction, ion-exchange and gel-filtration chromatography. The native and subunit molar mass was 330 and 110 kDa, respectively. The pH and temperature optima were 4.6-5.3 and 70 degrees C, respectively. The K(m) and kcat for 4-nitrophenyl beta-D-glucopyranoside at 40 degrees C and pH 5 were 1.11 mmol/L and 4000/min, respectively. The enzyme was activated by low and inhibited by high concentrations of NaCl. Ammonium sulfate inhibited the enzyme. Thermolysin periodically inhibited and activated the enzyme during the course of reaction and after 150 min of proteinase treatment only 10% activity was lost with concomitant degradation of the enzyme into ten low-molar-mass active bands. When subjected to 0-9 mol/L transverse urea-gradient-PAGE for 105 min at 12 degrees C, the nonpurified beta-glucosidase showed two major bands which denatured at 4 and 8 mol/L urea, respectively, with half-lives of 73 min.
Rashid
M H
MH
National Institute for Biotechnology and Genetic Engineering, Faisalabad, Pakistan.
Siddiqui
K S
KS
D000645:Ammonium Sulfate / Q000494:pharmacology; D001234:Aspergillus niger / Q000201:enzymology*; D011232:Chemical Precipitation; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005656:Fungal Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D013820:Thermolysin / Q000494:pharmacology; D001617:beta-Glucosidase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism
D004791:Enzyme Inhibitors; D005656:Fungal Proteins; D001617:beta-Glucosidase; D013820:Thermolysin; D000645:Ammonium Sulfate