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)

The study of the acidic lipid requirement of human spleen glucocerebrosidase was extended to include two new series of acidic lipids, namely, monoacylglycol sulfates and diacylglycerol sulfates. Lysosomal glucocerebrosidase was extracted with sodium cholate and 1-butanol to render its beta-glucosidase activity dependent upon exogenous lipids. Maximum reactivation of control glucocerebrosidase was obtained with nonanoylglycol sulfate (NGS) and diheptanoylglycerol sulfate (DHGS). However, the effects of these lipids were markedly dependent on the nature of buffer used in the assay medium; specifically, 0.2 M sodium citrate-phosphate (pH 5.5) was much more effective than 0.2 M sodium acetate (pH 5.5) in permitting these lipids to reactivate glucocerebrosidase. In contrast, the marked activation of glucocerebrosidase by phosphatidylserine and galactocerebroside 3-sulfate (sulfatide) that was achievable in the sodium acetate buffer was totally inhibited by citrate or phosphate ions. The effects of NGS and DHGS on the kinetic parameters of control glucocerebrosidase were to lower the Km for the substrate, 4-methylumbelliferyl-beta-D-glucoside from 5.5 mM to approximately 2 mM (in sodium citrate-phosphate buffer) and markedly increase the Vmax. Furthermore, with DHGS, significant activation was achieved at concentrations below the lipid's critical micellar concentration. None of the monoacylglycol- or diacylglycerol sulfates were capable of stimulating mutant glucocerebrosidases from either type 1 (Ashkenazi-Jewish) or type 2 Gaucher's disease patients. Like control glucocerebrosidase, the type 1 glucocerebrosidase was unresponsive to phosphatidylserine and sulfatide when the beta-glucosidase assay was conducted in 0.2 M sodium citrate-phosphate buffer. Based on the differential action of these lipid activators in the two buffers and their effects on the mutant enzymes, we propose that, with regard to the lipid requirement of glucocerebrosidase, there are two classes of acidic lipids--one comprised of phosphatidylserine and sulfatide and the other comprised of the likes of NGS, DHGS, or sodium taurodeoxycholate. It appears that control glucocerebrosidase and the mutant enzyme of the patient with type 1 Gaucher's disease is reconstitutable with the first class of lipids whereas the glucocerebrosidase of the type 2 patient is not. The observations in this report are interpreted in terms of a model which postulates that normal glucocerebrosidase possesses at least two distinct lipid binding domains.
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PMID:Activation of human spleen glucocerebrosidases by monoacylglycol sulfates and diacylglycerol sulfates. 335 73

We show that sialosylgangliotetraosylceramide (GM1) is a potent activator of delipidated (sodium cholate- and 1-butanol-extracted) lysosomal rat liver glucocerebroside:beta-glucosidase. Stimulation of 4-methylumbelliferyl-beta-D-glucopyranoside hydrolysis by the beta-glucosidase was markedly dependent upon the concentration of GM1 in the assay medium. Estimations of critical micellar concentration (CMC) performed fluorometrically using the dye N-phenylnaphthylamine revealed two CMC values of GM1 above 18 degrees C; the CMC of the primary micelles (3.32 microM) was temperature-independent whereas that of the secondary micelles decreased with decreasing temperature (17.2 and 10.8 microM at 37 and 20 degrees C, respectively). In the temperature range of 18-39 degrees C, beta-glucosidase activity increased sharply when the GM1 concentration was above the CMC of the secondary micelles. Although a heat-stable factor, purified from the spleen of a patient with Gaucher's disease, had a profound effect on the activation of beta-glucosidase by GM1, it decreased the CMC only slightly (14.8 versus 17.2 microM at 37 degrees C). The heat-stable factor (8 micrograms/ml) changed the shape of the activation curve from sigmoidal to hyperbolic, suggesting that the heat-stable factor permits beta-glucosidase to be activated by primary micelles or monomers. The results of gel filtration chromatography and sucrose gradient centrifugation in H2O and D2O revealed that the activation of beta-glucosidase by GM1 was associated with an increase in the size of the enzyme from 45,800 to 178,500 daltons and an increase in the partial specific volume from 0.697 to 0.740 ml/g. The active, reconstituted beta-glucosidase appears to consist of 50% protein and 50% ganglioside (56 molecules/178,500 g). Concentrations of GM1 below the CMC of secondary micelles increased the rate of inactivation of the enzyme by the irreversible inhibitor conduritol B epoxide at 37 degrees C, indicating that GM1 monomers or primary micelles do interact with the enzyme, even though they do not increase the rate of hydrolysis of 4-methylumbelliferyl-beta-D-glucopyranoside by the enzyme.
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PMID:Characterization of the activation of rat liver beta-glucosidase by sialosylgangliotetraosylceramide. 393 39

The acidic phospholipid requirement of the predominant particulate beta-glucosidase of mammalian spleen and liver was investigated using a series of N-acyl derivatives of dioleoyl phosphatidylethanolamine (PE). The PE, a neutral phospholipid, was converted to an acidic lipid, (N-acyl)-phosphatidylethanolamine (NAPE) by acylation of the amino group with different fatty acyl chains. Lysosomal beta-glucosidases from rat liver and spleens of controls and patients with various types of Gaucher's disease were solubilized and delipidated by extraction with sodium cholate and 1-butanol. All members of the NAPE series tested were effective activators of the delipidated rat liver beta-glucosidase, and the stimulatory power of the NAPE family increased with increasing chain length of the fatty acid substitution. In contrast, dioleoyl-PE had no effect on beta-glucosidase activity. A heat-stable factor (HSF) purified from the spleen of a patient with Gaucher's disease significantly increased the sensitivity of the rat liver beta-glucosidase to all of the NAPE derivatives. The maximum stimulation achieved in the presence of HSF was independent of N-acyl chain length. Compared to the potent activator, phosphatidylserine (PS), (N-acetyl)-PE and (N-linoleoyl)-PE were half as effective as activators of beta-glucosidase from control human spleen. PS stimulated the beta-glucosidase of type 1 nonneurologic Gaucher's disease, but none of the NAPE compounds activated it. Neither PS nor any of the (N-acyl)-PE compounds could activate a delipidated preparation of beta-glucosidase obtained from the spleen of a neurologic case. These results indicate that although the presence of a net negative charge on a phospholipid confers upon it an ability to reconstitute beta-glucosidase activity to the normal, nonmutant enzyme, it is insufficient to permit differentiation of the various types of Gaucher's disease.
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PMID:Comparison of N-acyl phosphatidylethanolamines with different N-acyl groups as activators of glucocerebrosidase in various forms of Gaucher's disease. 393 29

The lipid requirement of membrane-bound rat liver beta-glucosidase was investigated using 4-methylumbelliferyl-beta-D-glucopyranoside as the substrate. The enzyme was solubilized and delipidated by sequential extraction of a crude lysosomal fraction from rat liver lysosomes with sodium cholate and ice-cold butan-1-ol. Neither saturated nor unsaturated phosphatidylcholine activated this enzyme. In contrast, acidic phospholipids like phosphatidylglycerol (PtdGro) and phosphatidylserine (PtdSer) were effective activators. For the PtdGro series, fatty acid composition was important, with the shorter chain or unsaturated fatty acid-containing PtdGro species being the best activators. Heat-stable factor (HSF) from Gaucher spleen by itself (1-2 micrograms) had no effect on enzyme activity. However, the same amount of HSF when combined with 10 micrograms of PtdSer markedly stimulated beta-glucosidase activity. In the presence of HSF, di-9-cis-octadecenoyl-PtdGro (1 microgram) or -PtdSer (5 micrograms) provided maximum protection of beta-glucosidase against heat (60 degrees C) inactivation. In the absence of phospholipids, HSF had no effect on the rate of inactivation of the enzyme by the suicide inhibitor conduritol B epoxide (t0.5, 12 +/- 0.5 min); the maximum rate of inactivation was achieved in the presence of a mixture of PtdGro (2.5-5 micrograms) and HSF (t0.5, 2.8 min). The combination of PtdSer (10 micrograms) and HSF (1.3 micrograms) lowered the Km for 4-methylumbelliferyl-beta-D-glucopyranoside from 24 to 2.7 mM. Inhibition of the enzyme by the glucocerebrosidase substrate analogues N-hexyl-O-glucosylsphingosine and glucosylsphingosine was influenced by the activator substances. The inclusion of PtdSer and HSF in the beta-glucosidase assay medium lowered the Ki of N-hexyl-O-glucosylsphingosine 20-fold. The same combination of activators decreased the I0.5 of the enzyme for glucosylsphingosine from 89.4 to 7.6 microM. A study of log (Vmax./Km) versus pH indicated that the PtdSer-HSF pair creates the active site of beta-glucosidase, making apparent three ionizable groups on the enzyme with pK values in the range 4.5-5.1.
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PMID:Characterization of the phospholipid requirement of a rat liver beta-glucosidase. 651 62

Agrobacterium tumefaciens beta-glucosidase, Cbg1 was extensively characterised and found to be a retaining aryl-glucosidase and an aryl-xylosidase. Cbg1s specificity for p-nitrophenyl beta-d-xylopyranoside was 73% that for p-nitrophenyl beta-d-glucopyranoside when measured by the ratio kcat/Km. The enzyme also hydrolysed p-nitrophenyl beta-d-fucopyranoside, and p-nitrophenyl beta-d-galactopyranoside with moderate efficiency. The enzyme released only terminal glucose from p-nitrophenyl beta-cellobioside and had a 20 000-fold preference for its natural substrate coniferin over cellobiose as indicated by the ratio kcat/Km. The enzyme was activated in the presence of 20 mM 1-propanol, 1-butanol, 1-pentanol, 1-hexanol, 1-heptanol, and 1-octanol. In the case of butanol this activation was investigated and shown to be due to transglycosylation activity with over 80% of p-nitrophenyl beta-d-glucopyranoside being converted to 1-butyl beta-d-glucopyranoside in the presence of Cbg1 and 100 mM 1-butanol.
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PMID:Agrobacterium tumefaciens beta-glucosidase is also an effective beta-xylosidase, and has a high transglycosylation activity in the presence of alcohols. 963 May 31