Purification and characterization of beta-xylosidase from Aureobasidium pullulans

• Published in: Applied microbiology and biotechnology Vol. 35; no. 2; pp. 210 - 215
• Main Authors: Dobberstein, J, Emeis, C.C
• Format: Journal Article
• Language: English
• Published: Berlin Springer 01-05-1991
• Subjects: Aureobasidium pullulans; Biological and medical sciences; Biotechnology; enzyme activity; Enzyme engineering; Fundamental and applied biological sciences. Psychology; Improved methods for extraction and purification of enzymes; Methods. Procedures. Technologies; purification; xylan 1,4- beta -xylosidase; Purification; Enzyme; Molecular weight determination; Induction; Exo-1,4-β-D-xylosidase; Gel permeation chromatography; Ion exchange chromatography; Fungi; Enzymatic activity; Aureobasidium pullulans; Substrate specificity; Production; Fungi Imperfecti; Kinetic parameter; Thallophyta
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/BF00184688

beta-Xylosidase was obtained from Aureobasidium pullulans CBS 58475 with an activity of 0.35 units/ml culture filtrate. The production of the enzyme was strongly inducible. beta-Xylosidase was purified in two steps by anion exchange and gel-permeation chromatography to high purity. The enzyme is a glycoprotein with an apparent molecular mass of 224 kDa in sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and separates into two subunits of equal molecular mass. After SDS-PAGE beta-xylosidase could be renatured and stained with methylumbelliferyl-beta-xylopyranoside. The enzyme was able to split substrates of other glycosidases. The maximum activity was reached at pH 4.5 and 80 degrees C. beta-Xylosidase showed high stability over a broad pH range from pH 2.0 to 9.5 and up to 70 degrees C. Analysis of cleavage patterns revealed that the enzyme was a typical glycosidase. Larger oligosaccharides consisting of xylose were degraded by an exomechanism together with a transxylosylation reaction.