Gene/Protein Disease Symptom Drug Enzyme Compound
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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)

This enzyme shows beta-D-glucosidase, beta-D-fucosidase and beta-D-galactosidase activities, all associated in a single peak in Sephadex G-200, DEAE-cellulose, concanavalin A-Sepharose chromatographies, and in high resolution isoelectric focusing (pI 4.56), having the optimal pH in the range 4.5-5.5. The enzyme is very stable under different conditions: (i) at pH in the range 5.5-7.0; (ii) in successive freezing-thawing cycles; (iii) at 4 degrees C; (iv) after exhaustive ultrasonic treatment. It is not stable beyond 40 degrees C, and in the presence of urea, Triton X-100, SDS or mercaptoethanol. HgCl2, KCN, Tris, maltose and the lactones were inhibitors of the enzyme. With glucose, fucose and galactose the inhibition is competitive. In addition, a transglycosylation mechanism seems to occur. The kinetic studies suggest a substrate-activation model and the presence of two primary active sites: fuco-gluco and galacto.
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PMID:Characterization and kinetics of beta-D-gluco/fuco/galactosidase from sheep liver. 641 26

A fluorescent derivative of glucosyl ceramide was synthesized by covalently linking a fluorescent fatty acid, 12-[N-methyl-N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)] aminododecanoic acid to the amino group of sphingosyl-1-O-beta-D-glucoside, glucosyl sphingosine. For hydrolysis by glucocerebrosidase, this substrate was dispersed in mixed micelles with Triton X-100 and sodium taurocholate or in unilamellar liposomes with phosphatidylcholine and the negatively charged lipid, dicetylphosphate. In either micellar or liposomal dispersions of the fluorescent substrate, reaction rates were linear with time and protein concentration, and saturation kinetics were observed. The rate of hydrolysis of this fluorescent substrate was equal to that obtained with radiolabeled glucosyl ceramide. The fluorescent glucosyl ceramide was used to determine glucocerebrosidase activity in extracts of human leukocytes, cultured skin fibroblasts, and various tissues as well as in partially purified splenic and placental glucocerebrosidase preparations. This fluorescent derivative of the natural substrate was not hydrolyzed by aryl beta-glucosidase(s), thereby facilitating the specific and reliable diagnosis of heterozygotes and homozygotes with Gaucher disease.
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PMID:Synthesis of a fluorescent derivative of glucosyl ceramide for the sensitive determination of glucocerebrosidase activity. 642 2

Studies were undertaken to characterize the beta-glucosidase activity in freshly homogenized liver from Sprague-Dawley rats. About 95% of the total beta-glucosidase activity was associated with the particulate fraction, whereas only about 3-7% was found in the cytosol. Storage of fresh liver at room temperature for several hours or repeated freezing and thawing of fresh rat liver prior to homogenization, solubilized 20-30% of the total hepatic beta-glucosidase activity. An additional 30% could be solubilized by extracting the particulate sediments with water or Triton X-100. The enzymatic activity in both the particulate and solubilized fractions optimally hydrolyzed 4-methylumbelliferyl-beta-D-glucoside as well as the glycolipid substrate, glucosylceramide, at an acidic pH. The rates of hydrolysis of either substrate by all subcellular fractions were stimulated by addition of sodium taurocholate or phosphatidylserine. The particulate, cytosolic and solubilized enzymes bound to concanavalin A, were inhibited by conduritol B epoxide and migrated more electronegatively on cellulose acetate than the cytosolic acid beta-glucosidase from human liver or spleen. These data indicated that the liver of Sprague-Dawley rats contained primarily the lysosomal acid beta-glucosidase ('glucocerebrosidase') and little, if any, 'nonspecific' beta-glucosidase. This, and the fact that about 60% of the rat hepatic beta-glucosidase could be solubilized by autolysis, freezing and rethawing or extraction with water, contrasts with the beta-glucosidases in human liver since about 80% of the total beta-glucosidase activity is cytosolic and does not hydrolyze glucosylceramide.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Lysosomal beta-glucosidase of rat liver. 642 48

Studies have been carried out on the activities and properties of the isozymes of alpha-mannosidase, alpha-glucosidase and beta-glucosidase in granulocytes, monocytes, lymphocytes and platelts from peripheral blood of heatlhy adult donors. The findings reveal the differences in activities as well as a characteristic distribution of the different molecular forms of these lysosomal hydrolases in specific cell types. Therefore, the results obtained with unfractionated total leukocyte smples from different subjects may vary according to the distribution of cell types in the circulation. Granulocytes and monocytes show only the acid alpha-mannosidase activity whereas lymphocytes and platelets show both acid and neutral activities. The specific activity of acid alpha-mannosidase in granulocytes and monocytes is higher than in lymphocytes and platelets. By DEAE-cellulose chromatography, the acid alpha-mannosidase in granulocyte and monocyte extracts elutes as two peaks, but only one peak is seen in lymphocytes. All cell types show both acid and neutral alpha-glucosidase activities. The specific activities of both isozymes are higher in granulocytes and monocytes than in lymphocytes and platelets. Monocytes show a higher acid than neutral activity. All other cell types show a higher neutral activity. Beta-Glucosidase in all cell types is mainly membrane-bound and it can be released by Triton X-100 and sodium taurocholate. Taurocholate also stimulates the beta-glucosidase activity of granulocytes, monocytes and lymphocytes whereas it inhibits the activity of this enzyme in platelets. These results indicate that variations in the total number of leukocytes and in the relative proportion of the various cell types in health and disease may yield inconsistent or unreliable values for enzyme activity in the diagnosis of lysosomal storage disease and in carrier detection.
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PMID:Studies on the activities and properties of lysosomal hydrolases in fractionated populations of human peripheral blood cells. 676 26

A reproducible and convenient method for assaying glucocerebrosidase activity using the natural substrates has been developed. From the insoluble pellet fraction of cultured skin fibroblast homogenates, released glucose was measured enzymically using hexokinase coupled with the glucose-6-phosphate dehydrogenase (G6PD) and nicotinamide adenine dinucleotide phosphate (NADP) system. Optimal enzyme assay conditions required both Triton X-100 and sodium taurocholate, pH 4.8. Glucocerebrosidase activities from three patients with type 1 Gaucher disease were 17.5%, 15.8%, and 11.2% of normal (normal = 198 +/- 14 nmol/hr per mg protein, n = 3). The first patient had normal beta-glucosidase activity with the artificial fluorogenic umbelliferone substrate. Interference with the accuracy of the glucose-dependent assay system by either glycolytic or gluconeogenic enzyme activites was not detected under these experimental conditions, and when substrates with long fatty-acid chain lengths (C = 22) were used, markedly decreased glucocerebrosidase activity occurred in both normal individuals and patients. The apparent Km's for the natural substrates were 0.56 +/- 0.05 mM for controls and 2.2-3.3 mM for Gaucher fibroblasts. These data further support the hypothesis that a structurally altered and catalytically deficient enzyme is synthesized in patients with type 1 Gaucher disease and illustrate the value of the natural substrate in investigating patients.
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PMID:Gaucher disease. III. Substrate specificity of glucocerebrosidase and the use of nonlabeled natural substrates for the investigation of patients. 677 30

We have previously reported a decreased activity of the lysosomal enzyme dipeptidyl aminopeptidase-I (DAP-I) in cultured fibroblasts from patients with Duchenne's muscular dystrophy (DMD). Here we report that electron microscope examination of these cells reveals the presence of abundant lamellar bodies, a morphologic abnormalities commonly associated with impaired lysosomal function. Morphometric analysis of these cytoplasmic figures in dystrophic cells shows a sevenfold increase relative to normal controls (P less than 0.01). Analysis of lysosomal density profiles by density gradient centrifugation reveals similar patterns in normal and DMD cells. Treatment of lysosomes wit the nonionic detergent Triton X-100 causes an activation of DAP-I. This activation, attributable to structure-linked latency, is markedly diminished in DMD cells which show an optimal activation of only 180% compared to 255% for control fibroblasts (P less than 0.01). These data suggest an alteration in the properties of the lysosomal membrane in DMD fibroblasts. This suggestion is also supported by studies on the release of DAP-I from lysosomes by osmotic shock which show it to be a membrane-associated enzyme with membrane-binding characteristics intermediate between those of tightly bound beta-glucosidase and those of unbound N-acetylgalactosaminidase. The latency characteristics of these other lysosomal enzymes are not altered in the DMD cells, indicating that the effect is specific for DAP-I.
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PMID:Structural changes in lysosomes from cultured human fibroblasts in Duchenne's muscular dystrophy. 678 12

1. A third beta-glucosidase from human liver has been isolated using a mild (0.02-0.10%) Triton X-100 extraction of the exhaustively washed high speed (200,000 X g, 30 min) particulate fraction, QAE-Sephadex and concanavalin A-Sepharose chromatography. This new beta-glucosidase, referred to as TX beta-glucosidase, possesses a distinctive set of chemical properties such that it is similar to both, glucocerebrosidase and cytoplasmic beta-glucosidase, but it is not identical to either enzyme. 2. The TX beta-glucosidase hydrolyzes glucocerebroside as well as the beta-D-glucose, beta-D-galactose, beta-D-fucose, beta-D-xylose and alpha-L-arabinose derivatives of 4-methylumbelliferone. Like the cytoplasmic beta-glucosidase, the TX beta-glucosidase is inhibited by bile salts, and unaffected by conduritol B epoxide and heat stable activator protein. 3. All three beta-glucosidases were inhibited by N-hexylpsychosine, and all showed the same, mixed type inhibition kinetics, indicating a common hydrophobic binding site in all three enzymes. 4. The TX beta-glucosidase, which constitutes only a few percent of the total beta-glucosidase activity of human liver, is absent from liver from two cases of neurologic Gaucher disease and present in reduced amounts in a third case with CNS disease. Liver from a case of type 1 Gaucher disease contained normal amounts of the TX beta-glucosidase.
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PMID:Effectors of three beta-glucosidases from human liver. 681 77

Glucosidase activities capable of removing the three glucose residues from Glc3Man9GlcNAc2 oligosaccharide were detected in a cell-free preparation of Saccharomyces cerevisiae X-2180. The glucosidase which cleaves the glucose residue at the nonreducing terminus (Glc3Man9GlcNAc2 oligosaccharide glucosidase) was equally distributed between the particulate and the supernatant fractions obtained after centrifugation of the yeast homogenate at 27,000 X g for 30 min. The membrane-bound activity was stimulated by Triton X-100, whereas the supernatant activity was not affected. The soluble Glc3Man9GlcNAc2 oligosaccharide glucosidase was partially purified from the supernatant by ammonium sulfate fractionation followed by DEAE-Sephadex chromatography. It was clearly separated from alpha-glucosidase, which acts onp-nitrophenyl-alpha-D-glucopyranoside, but still contained beta-glucosidase and alpha-mannosidase acting on p-nitrophenyl-beta-D-glucopyranoside and alpha-D-mannopyranoside, respectively. The Glc3Man9GlcNAc2 oligosaccharide glucosidase had a pH optimum of 6.8, and showed no requirement for divalent cations. The enzyme was very active with glucose-labeled Glc3Man9GlcNAc2, was slightly active with Glc2Man9GlcNAc2, and showed no activity with Glc1Man9GlcNAc2. These properties suggest that this enzyme is involved in the first step of processing of oligosaccharides after transfer from dolichyl pyrophosphate to proteins.
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PMID:Partial purification from Saccharomyces cerevisiae of a soluble glucosidase which removes the terminal glucose from the oligosaccharide Glc3Man9GlcNAc2. 701 69

Glycosidases and glycosyltransferases were electrophoresed in the presence of sodium dodecyl sulfate (SDS) in a thin-layer gel supported by a glass plate, treated with the nonionic detergent Triton X-100, and specifically stained for the sugar-releasing activity of these enzymes. Staining is based on conversion of monosugars or a sugar phosphate to glucose-6-phosphate by the appropriate intermediary enzymes, reduction of NADP+ to NADPH, and accumulation of reduced Nitroblue Tetrazolium in the gel. Among the enzymes tested, alpha-glucosidase, beta-glucosidase and beta-mannosidase could not be renatured, whereas beta-fructofuranosidase and alpha-mannosidase could be renatured unless heated before electrophoresis. Sucrose phosphorylase, glucosyltransferase and fructosyltransferase, which are single-peptide proteins with no cystine bond, could be renatured even after pretreatment with SDS and/or mercaptoethanol at 100 degrees C for 10 min. However, exclusive heating remarkably decreased the activities of these enzymes. Two-dimensional separation of the five renaturable enzymes was done in a single thin-layer gel, using SDS-electrophoresis in the first dimension and isoelectric focusing in the second dimension.
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PMID:Renaturation and activity staining of glycosidases and glycosyltransferases in gels after sodium dodecyl sulfate-electrophoresis. 752 70

The production of beta-glucosidase by Aspergillus terreus was investigated in liquid shake cultures. Enzyme production was maximum on the 7th day of growth (2.18 U/ml) with the initial pH of the medium in the range of 4.0-5.5. Cellulose (Sigmacell Type 100) at 1.0% (wt/vol) gave maximum beta-glucosidase activity among the various soluble and insoluble carbon sources tested. Potassium nitrate was a suitable nitrogen source for enzyme production. Triton X-100 at 0.15% (vol/vol) increased the enzyme levels of A. terreus. The test fungal strain showed an ability to ferment glucose to ethanol.
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PMID:Production of beta-glucosidase by Aspergillus terreus. 776 52


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