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Query: EC:3.2.1.21 (beta-glucosidase)
 3,280 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Although beta-D-fucosidase (beta-D-fucoside fucohydrolase, EC 3.2.1.38) has been isolated from various sources, the identity of this enzyme is still not settled. We have purified a specific beta-D-fucosidase in electrophoretically homogeneous form crude extracts of Aspergillus phoenicis by polyethyleneglycol 6000-phosphate buffer aqueous two-phase separation, and successive chromatography on DEAE-Sephadex A-50, hydroxyapatite and Sephadex G-100 columns. The molecular weight of the enzyme was estimated to be 57000 by SDS-polyacrylamide gel electrophoresis and 50000 to 60000 by gel filtration on Sephadex G-100. The enzyme showed optimum coside were 2.4mmol/L, and 1.28 mumol min-1 the pH range 5.5-6.5 and below 35 degrees C. The Km and the Vmax values for pNP-beta-D-fucoside were 2.4mmol/L, and 1.28 mumol.min-1.mg-1 respectively. The enzyme was strongly inhibited by sulfhydryl group reagents, PCMB-NEM and iodoacetate. It was also inhibited by EDC, DEP and NBS. Thus, -SH, -COOH groups, histidyl and tryptophyl residues were essential for enzyme activity. The purified beta-D-fucosidase showed high specificity toward p-nitrophenyl beta-D-fucoside. The enzyme was inhibited by D-fucose and D-fucono-gamma-lactone, but not by D-galactose, D-galactono-gamma-lactone, D-glucose or D-glucono-gamma-lactone; the latter compounds are specific inhibitors of beta-D-galactosidase and beta-D-glucosidase respectively. Thus, this enzyme is the most strictly specific beta-D-fucosidase when compared with those previously reported.
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PMID:[Studies on the beta-D-fucosidase from Aspergillus phoenicis]. 159 57

Purified beta-glucosidase from Aspergillus niger NIAB280 was chemically modified by l-ethyl-3-(3-dimethyl-aminopropyl)-carbodi-imide (EDC) in the presence of glycinamide (GAM) as nucleophile under various conditions to study the role of carboxy groups in the catalytic mechanism of this enzyme. beta-Glucosidase was inactivated by the binding of one mol of EDC per mol of the enzyme with a second-order rate constant of 4.77 x 10(-2) mM min-1. Glucose, as competitive inhibitor, partly protected the active-site carboxy group against chemical modification, with a Kd of 3.64 mM. The pH dependence of chemical modification by EDC showed that first-order rate constants decreased with increasing pH, indicating that the proton donating group is a carboxy group. The pKa values of the acidic and basic limbs of the native enzyme were 2.9 and 6.5 respectively. beta-Glucosidase was modified by EDC in the presence of GAM and ethylenediamine dihydrochloride (EDAM) as nucleophiles for 60 min. The effects of neutralization (GAM) and reversal (EDAM) of the negative charges of surface carboxy groups on the kinetic properties of the enzyme were also studied. Native beta-glucosidase, GAM and EDAM had Vmax/K(m) values of 0.73, 1.22 and 0.60 respectively at 40 degrees C. Interestingly, the activation energy profiles of native beta-glucosidase (103 and 79 kJ/mol) were biphasic, whereas those of GAM (137, 101 and 30 kJ/mol) and EDAM (285, 100 and 29 kJ/mol) were triphasic, indicating significant activation of modified beta-glucosidases at temperatures higher than 50 degrees C. The pKa values of both the active-site carboxy groups as well as the pH optima of GAM and EDAM were also significantly decreased compared with those of the native beta-glucosidase.
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PMID:Carboxy-group modification: high-temperature activation of charge-neutralized and charge-reversed beta-glucosidases from Aspergillus niger. 966 79