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A novel and direct assay for measuring enzymatic depolymerisation of β‐1,3‐glucans
Author(s) -
Aktas Lale Y.,
Adlard Max,
Trevan Michael,
Guven Avni,
Brownleader Michael D.
Publication year - 2000
Publication title -
phytochemical analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.574
H-Index - 72
eISSN - 1099-1565
pISSN - 0958-0344
DOI - 10.1002/1099-1565(200009/10)11:5<301::aid-pca518>3.0.co;2-6
Subject(s) - chemistry , glucanase , detection limit , enzyme , glucan , hydrolysis , chromatography , polymer , biochemistry , organic chemistry
Indirect colorimetric analyses of released reducing groups, as an indicator of induced β‐1,3‐glucanase activities, tend to be the assay methods of choice for the characterization of plant endo‐ and exo‐β‐1,3‐glucanase. However, interfering low molecular weight reducing sugars found in unprocessed plant extracts often mask in vitro estimations of β‐1,3‐glucanase activity. The enzyme‐catalysed hydrolysis of an optically active β‐1,3‐glucan polymer was monitored polarimetrically by measuring changes in the rotation of plane‐polarized light of the reaction assay medium. A direct, non‐radiochemical assay that detects changes in the optical rotation of released (1→3) β‐ D ‐oligoglucosides with low degrees of polymerisation (≪25) has been found to be highly reproducible, rapid (total analysis time 15 min), sensitive (detection limit 0.007 unit/mL glucanase), selective and non‐destructive. This assay has been used to detect a salicylate‐induced cell wall‐bound tomato β‐1,3‐glucanase, which is a component of a pathogenesis‐related response in plants. Copyright © 2000 John Wiley & Sons, Ltd.