Partial Purification and Properties of an Induced β‐d‐Xylosidase of Bacillus pumilus 12

Abstract

A highly specific β‐d‐xylosidase with absolute glycon‐substrate specificity was induced in Bacillus pumilus 12 by xylose. It was purified 34‐fold by a procedure involving lysis of the cells with lysozyme, ammonium sulfate fractionation, G‐75 Sephadex gel filtration and hydroxylapatite chromatography. The optimal activity was in the pH region 7.0–7.3. The isoelectric point was found to be 4.4. In contrast to its broad complexing affinity, the hydrolytic activity was restricted to aryl β‐d‐xylopyranosides and lower xylo‐oligosaccharide derivatives. From the dependence of Km and Vmax on pH, it could be deduced that two dissociable groups were necessary for catalytic activity. The enzyme was competitively inhibited by Tris at pH 7.2 and proved sensitive to ionic environment. p‐Chloromercuribenzoate and other SH‐reagents were inhibitory but substrate analogs and competitive inhibitors protected the enzyme against inactivation by p‐chloromercuribenzoate. The results indicate that a histidine residue or imidazolium group and a sulf‐hydryl group might participate in the enzymatic hydrolysis of p‐nitrophenyl β‐d‐xylopyranoside. Copyright © 1969, Wiley Blackwell. All rights reserved