PURIFICATION AND CHARACTERIZATION OF A GLUCOSE-TOLERANT BETA-GLUCOSIDASE FROM ASPERGILLUS-NIGER CCRC-31494

An extracellular glucose-tolerant beta-glucosidase was purified to hom ogeneity by alcohol fractionation and preparative isoelectric focusing from Aspergillus niger CCRC 31494, The enzyme was a dimeric protein w ith a subunit of 49,000, and had its optimum activity at pH 5.0 and 55 degrees C, The enzyme was completely inhibited by 5 mM Ag+, Thiol gro ups and serine residues were not essential for its activity, Low conce ntrations of alcohols (10%) except for methanol could activate the enz yme, It was very specific for para-nitrophenyl-beta-D-glucoside (pNPG) and cellobiose. However, the enzyme also had some beta-xylosidase act ivity, but showed no activity towards alpha-linked glycosidic substrat es, The V-max of 124.4 U/mg and 21.6 U/mg were found for pNPG (K-m=21. 7 mM) and para-nitrophenyl-beta-D-xyloside (pNPX) (K-m=14.2 mM), respe ctively, The enzyme was tolerant to glucose inhibition with a K-i of 5 43 mM, while fructose, galactose, mannose, and xylose were not inhibit ory.