PURIFICATION AND CHARACTERIZATION OF AN EXTRACELLULAR BETA-GLUCOSIDASE WITH TRANSGLYCOSYLATION AND EXO-GLUCOSIDASE ACTIVITIES FROM FUSARIUM-OXYSPORUM

An extracellular beta-glucosidase from Fusarium oxysporum was purified to homogeneity by gel filtration and ion-exchange chromatographies. T he enzyme, a monomeric protein of 110 kDa, was maximally active at pH 5.0-6.0 and at 60 degrees C. It hydrolysed 1-->4-linked aryl-beta-gluc osides and 1-->4-linked, 1-->3-linked and 1-->6-linked beta-glucosides . The apparent K-m and k(cat) values for p-nitrophenyl beta-D-glucopyr anoside (4-NpGlcp) and cellobiose were 0.093 (K-m), 1.07 mM (k(cat)) a nd 1802 (K-m), 461.5 min(-1) (k(cat)), respectively. Glucose and gluco nolactone inhibited the enzyme competitively with K-i values of 2.05 m M and 3.03 mu M, respectively. Alcohols activated the enzyme; butanol showed maximum effect (2.2-fold at 0.5 M) while methanol increased the activity by 1.4-fold at 1 M. The enzyme catalysed the synthesis of me thylglucosides, ethylglucoside and propylglucosides, as well as trisac charides in the presence of different alcohols and disaccharides, resp ectively In addition, the enzyme hydrolysed the unsubstituted and meth ylumbelliferyl cello-oligosaccharides [MeUmb(Glc)(n)] but the rate of hydrolysis decreased with increasing chain length. Analysis of prod uc ts released from MeUmb(Glc)(n) as a function of time revealed that the enzyme attacked these substrates in a stepwise manner and from both e nds. Thus, beta-glucosidase from F. oxysporum, with the above interest ing properties, could be of commercial interest.