A Chromogenic substrate for a β-xylosidase-coupled assay of α-glucuronidase
Date
2000-08
Authors
Biely, Peter
Hirsch, Jan
La Grange, Daniel C.
Van Zyl, Willem H.
Prior, Bernard A.
Journal Title
Journal ISSN
Volume Title
Publisher
Elsevier
Abstract
-Nitrophenyl 2-(4-O-methyl-α- -glucopyranuronosyl)-β- -xylopyranoside obtained on deesterification of 4-nitrophenyl 2-O-(methyl 4-O-methyl-α- -glucopyranosyluronate)-β- -xylopyranoside (Hirsch et al., Carbohydr. Res. 310, 145–149, 1998) was found to be an excellent substrate for the measurement of hemicellulolytic α-glucuronidase activity. A new precise α-glucuronidase assay was developed by coupling the α-glucuronidase-catalyzed formation of 4-nitrophenyl β- -xylopyranoside with its efficient hydrolysis by β-xylosidase. A recombinant strain of Saccharomyces cerevisiae, harboring and expressing the β-xylosidase gene xlnD of Aspergillus niger under control of the alcohol dehydrogenase II promoter on a multicopy plasmid, was used as a source of β-xylosidase. The activity values of β-xylosidase in the assay required to achieve a steady-state rate of 4-nitrophenol formation shortly after starting the α-glucuronidase reaction were obtained both experimentally and by calculation using the kinetics of coupled enzyme reactions.
Description
The original publication is available at www.elsevier.com.
Includes bibliography.
Includes bibliography.
Keywords
α-Glucuronidase, Xylosidase, Enzymes, Microbiological assay, Chromogenic substrate, Glycoside, Aldobiuronic acid, Recombinant
Citation
Biely, Peter, Hirsch, Jan, La Grange, Daniel C., Van Zyl, Willem H., Prior, Bernard A. 2000. A Chromogenic Substrate for a b-Xylosidase-Coupled
Assay of a-Glucuronidase. Analytical Biochemistry 286:289-294.doi:10.1006/abio.2000.4810, http://www.sciencedirect.com.ez.sun.ac.za/science?_ob=MImg&_imagekey=B6W9V-45FK4RH-2H-3&_cdi=6692&_user=613892&_pii=S0003269700948103&_origin=search&_coverDate=11%2F15%2F2000&_sk=997139997&view=c&wchp=dGLbVtb-zSkzV&md5=97a7d694592b628ed93c14a774164170&ie=/sdarticle.pdf