Title: Characterization of an isozyme of beta-glucosidase from sweet almond
Authors: Li, YK
Chang, LF
Shu, HH
Chir, J
National Chiao Tung University
Department of Applied Chemistry
Keywords: sweet almond beta-glucosidase;beta-galactosidase;beta-D-glycopyranosides;isozyme;substrate specificity
Issue Date: 1-Feb-1997
Abstract: A sweet almond beta-glucosidase (EC isozyme was purified from commercial crude product. The process of purification consisted of a Protein-Pak Q anion exchange chromatography following by a Superdex 75 HR gel filtration separation. The purified enzyme is a monomeric glycoprotein with molecular weight of 58 kDa and pI = 4.55 which is distinguished from reported isozymes. The enzyme has a pH optimum in the range of 5.2-5.6 when p-nitrophenyl-beta-D-glycopyranosides are used as substrate and is stable up to 50 degrees C at that pH range. The purified protein also exhibits profound beta-galactosidase and alpha-L-arabinosidase activity. The study of substrate specificity revealed that lacking of hydroxymethyl group at C-5 of glycosides resulted in higher affinity for substrate binding to enzyme, whereas the chemical step of hydrolysis (k(cat)) was prevented significantly. The pH activity profile displayed a bell-shaped curve for all measured p-nitrophenyl-beta-D-glycopyranosides with apparent pK(1) and pK(2) values of 4.4-4.7 and 6.2-6.4, respectively. This isozyme was strongly inhibited by delta-gluconolactone (K-i = 160 mu M) and 4-phenylimidazole (K-i = 17.8 mu M) reversibly at pH 6.2. Among the tested glycoses, the binding affinity of N-acetyl-beta-D-glucosamine to the enzyme (K-i = 52 mM) was 6 times stronger than that of glucose and its epimers.
URI: http://hdl.handle.net/11536/761
ISSN: 0009-4536
Volume: 44
Issue: 1
Begin Page: 81
End Page: 87
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