Purification and some properties of Chaetomium trilaterale .BETA.-xylosidase.

Abstract

A β-xyloside hydrolytic enzyme of the fungus Chaetomium trilaterale was further purified by a modification of Kawaminami's procedure (DEAE-Sephadex A-25 and Sephadex G-75 column chromatography), followed by isoelectric focusing. The purified preparation was homogeneous by polyacrylamide disc gel electrophoreses at pH 4.3 and pH 8.3. The purified enzyme hydrolyzed β-D-glucopyranosides as well as β-D-xylopyranosides, and the ratio of β-glucosidase activity against β-xylosidase activity increased about 3 fold during the purification steps. The molecular weight of this preparation was estimated to be about 240, 000 by Sephadex G-200 gel filtration and 118, 000 by SDS-polyacrylamide slab gel electrophoresis. The isoelectric point was 4.86 and the amino acid composition was also determined. The optimum pH was at 4.2 for phenyl β-D-glucoside and around 4.5 for phenyl β-D-xyloside. The β-xylosidase activity was relatively stable but β-glucosidase activity was rapidly inactivated, at the alkaline pH range above 11. The heating of the preparation at 60°C didn't show a parallel inactivation of the two activities. N-Bromosuccinimide strongly inactivated both enzyme activities. Nojirimycin and glucono-l, 5-lactone showed a stronger inhibition on β-xylosidase activity than on β-glucosidase activity. The maximal velocities decreased in the order; phenyl β-D-glucoside > cellobiose > phenyl β-D-xyloside > xylobiose; the value with phenyl β-D-glucoside was about 28-fold higher than that with phenyl β-D-xyloside.