Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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 marine rotifer, Brachionus plicatilis, is able to digest Chlorella efficiently, suggesting that the rotifer contains a powerful cellulolytic enzyme system. A multi-component cellulolytic complex, including endoglucanase (CM-cellulase), cellobiohydrolase and beta-glucosidase, was found in Brachionus plicatilis. Endoglucanase (endo-beta-1,4 glucanase) was purified to homogeneity from rotifer homogenates using a sequential chromatographic method. The purified enzyme exhibits a strong hydrolytic activity with carboxymethyl(CM)-cellulose. The optimum temperature and pH for the endoglucanase activity were 37 degrees C and 7.0, respectively. 80% of the CM-cellulase activity was retained in salt mixture that ranged from 150 to 500 mM NaCl equivalent. The purified protein was isolated with a molecular weight of approximately 62 kDa estimated by SDS-polyacrylamide gel electrophoresis.
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PMID:Purification and characterization of an endoglucanase from the marine rotifer, Brachionus plicatilis. 935 Mar 31

During studies of the nutritional utilization of pyridoxine 5'-beta-D-glucoside, a major form of vitamin B6 in plants, we detected two cytosolic beta-glucosidases in jejunal mucosa. As expected, one was broad specificity beta-glucosidase that hydrolyzed aryl beta-D-glycosides but not pyridoxine beta-D-glucoside. We also found a previously unknown enzyme, designated pyridoxine-beta-D-glucoside hydrolase, that efficiently hydrolyzed pyridoxine beta-D-glucoside. These were separated and purified as follows: broad specificity beta-glucosidase 1460-fold and pyridoxine-beta-D-glucoside hydrolase 36,500-fold. Purified pyridoxine-beta-D-glucoside hydrolase did not hydrolyze any of the aryl glycosides tested but did hydrolyze cellobiose and lactose. Pyridoxine-beta-D-glucoside hydrolase exhibited a pH optimum of 5.5 and apparent molecular mass of 130 kDa by SDS-polyacrylamide gel electrophoresis and 160 kDa by nondenaturing gel filtration, in contrast to 60 kDa for native and denatured broad specificity beta-glucosidase. Glucono-delta-lactone was a strong inhibitor of both enzymes. Ionic and nonionic detergents were inhibitory for each enzyme. Conduritol B epoxide, a potent inhibitor of lysosomal acid beta-glucosidase, inhibited pyridoxine-beta-D-glucoside hydrolase but not broad specificity beta-glucosidase, but both were inhibited by the mechanism-based inhibitor 2-deoxy-2-fluoro-beta-D-glucosyl fluoride. Our findings indicate major differences between these two cytosolic beta-glucosidases. Studies addressing the role of vitamin B6 nutrition in regulating the activity and its consequences regarding pyridoxine glucoside bioavailability are in progress.
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PMID:Cytosolic pyridoxine-beta-D-glucoside hydrolase from porcine jejunal mucosa. Purification, properties, and comparison with broad specificity beta-glucosidase. 940 96

Carboxymethylcellulase (CMCase) was extracted and purified from an angiosperm parasite Cuscuta reflexa free from beta-glucosidase and other enzyme activities. The molecular mass and Stokes' radius of the purified enzyme are 144 kDa and 44 A, respectively. The diffusion coefficient and frictional ratio of the enzyme were 5.15 x 10(-7) cm2/sec and 1.27. The SDS-PAGE revealed homotetrameric nature of the enzyme with a subunit molecular mass of 35 +/- 1 kDa. Titration against DTNB and NBS revealed 19 sulfhydryl groups and 8 tryptophan groups, respectively, per mole of the enzyme. A sharp pH optimum at 5.0 was obtained. Cuscuta CMCase activity is unique amongst plant endoglucanases in being stimulated by Mg2+ and Mn2+ ions and by various thiols. Reaction product analysis, mode of enzyme action and substrate specificity test suggest the endo- nature of the purified CMCase. The enzyme showed K(m) value of 26 +/- 1 mg/ml for carboxymethylcellulose (sodium salt).
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PMID:Physico-chemical and functional characterization of a high molecular weight carboxymethylcellulase from Cuscuta reflexa. 949 45

A 4.7-kb DNA insert encoding a secretory cellobiase (Cba) was cloned from Cellulomonas biazotea in Escherichia coli using an excretion vector, pM. Host cells transformed with the recombinant construct, designated pBZ4.7, were able to utilize cellobiose as the sole carbon source. Part of the Cba activity encoded by pBZ4.7 could be detected in the periplasm and even in the culture supernatant. The Cba protein was purified from the culture supernatant and analyzed by SDS-PAGE to have an apparent M(r) of 86,000. The insert consisted of two PstI fragments with lengths of 0.75 and 3.95 kb, both of which were found to be crucial for expressing the Cba activity. Sequencing of the first 3.95 kb of the insert revealed that the coding sequence for Cba, designated the cba gene, was 2484 bp long. Comparison of the deduced Cba sequence with those of published beta-glucosidases revealed a potential active site located at the N-terminal portion of the former. The cba gene has a high G + C content of 76.4% and is flanked by a putative ribosome-binding site and potential transcriptional termination signals upstream and downstream from its coding sequence, respectively.
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PMID:The cloning, expression and characterization of a cellobiase gene encoding a secretory enzyme from Cellulomonas biazotea. 951 46

A large portion of beta-glucosidase (EC 3.2.1.21) in germinating rice seeds, which appears to be ionically bound to cell walls, can be solubilized with 1 M NaCl. Its activity increased more than eight-fold within five days of germination. It was purified to electrophoretic homogeneity from the extracts of germinated rice seeds by fractionation with (NH4)2SO4 followed by CM-Sepharose, Polybuffer exchanger 118, Concanavalin A-Sepharose and Bio-Gel P-100. The Mr of the purified enzyme, estimated by SDS-PAGE, was 56,000 and the isoelectric point was > 10.0. Its N-terminal amino acid sequence (44 residues) exhibited high homology to those of beta-glucosidases from other plants, such as barley and white clover. Its activity was optimal at pH 4.5 and 50 degrees, and it was strongly inhibited by glucono-1,5-lactone. The enzyme showed hydrolytic as well as transglycosylation activity towards (1-->3)-beta- and (1-->4)-beta-linked oligosaccharides with degree of polymerization of 2-4. The results suggest that the beta-glucosidase is probably involved not only in hydrolysis but also in modification of oligosaccharides in cell walls of germinating rice seeds.
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PMID:A cell wall-bound beta-glucosidase from germinated rice: purification and properties. 962 52

In plants, the naphthoquinone juglone is known to be involved in pathogenic defence mechanisms, but it may also take part in plant developmental processes. This naphthoquinone can accumulate in a glycosylated form, namely hydrojuglone beta-d-glucopyranoside. The structural configuration of this compound was shown to be 1, 5-dihydroxy-4-naphthalenyl-beta-d-glucopyranoside by means of MS, NMR and nuclear Overhauser effect spectroscopy analyses. A hydrojuglone beta-d-glucopyranoside beta-glucosidase (EC 3.2.1.21) was purified to homogeneity from Juglans regia L. The enzyme catalysed the release of juglone from hydrojuglone beta-d-glucopyranoside with high specificity and showed Michaelis-Menten kinetics with Km=0.62 mM and Vmax=14.5 microkat/mg of protein. This enzyme also showed a higher activity towards beta-d-fucosyl than beta-d-glucosyl bonds. The purified enzyme had an apparent Mr of 64000 by SDS/PAGE and a pI 8.9 by isoelectrofocusing PAGE. The purified enzyme was inhibited by several bivalent cations, such as Cu2+, Fe2+, Hg2+, and by d-glucono-1,5-lactone, showing non-competitive inhibition of the mixed type.
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PMID:Insight into naphthoquinone metabolism: beta-glucosidase-catalysed hydrolysis of hydrojuglone beta-D-glucopyranoside. 965 66

A cellobiohydrolase I (cbhI) and a beta-glucosidase 1 (bgl1) gene of Aspergillus aculeatus were expressed in Saccharomyces cerevisiae. The transformed cells secreted the enzymes efficiently in an active form. The recombinant CBHI gave two bands of different molecular mass (110 and 90 kDa) and the recombinant BGL1 gave one band (180 kDa) by SDS-PAGE. The recombinant CBHI and BGL1 had the same enzymatical properties as the native enzyme except for the specific activity toward cellulosic substrates. By the combination of three different types of cellulases, FI-CMCase, CBHI, and BGL1, we could hydrolyze Avicel up to 59% under our experimental conditions.
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PMID:Expression of Aspergillus aculeatus No. F-50 cellobiohydrolase I (cbhI) and beta-glucosidase 1 (bgl1) genes by Saccharomyces cerevisiae. 975 70

Plant cell suspension cultures of Rauwolfia produce within 1 week approximately 250 nkat/l of raucaffricine-O-beta-D-glucosidase. A five step procedure using anion exchange chromatography, chromatography on hydroxylapatite, gel filtration and FPLC-chromatography on Mono Q and Mono P delivered in a yield of 0.9% approximately 1200-fold enriched glucosidase. A short protocol employing DEAE sepharose, TSK 55 S gel chromatography and purification on Mono Q gave a 5% recovery of glucosidase which was 340-fold enriched. SDS-PAGE showed a Mr for the enzyme of 61 kDa. The enzyme is not glycosylated. Structural investigation of the enzyme product, vomilenine, demonstrated that the alkaloid exists in aqueous solutions in an equilibrium of 21(R)- and 21(S)-vomilenine in a ratio of 3.4:1. Proteolysis of the pure enzyme with endoproteinase Lys C revealed six peptide fragments with 6-24 amino acids which were sequenced. The two largest fragments showed sequences, of which the motif Val-Thr-Glu-Asn-Gly is typical for beta-glucosidases. Sequence alignment of these fragments demonstrated high homologies to linamarase from Manihot esculenta (81% identity) or to beta-glucosidase from Prunus avium (79% identity). Raucaffricine-O-beta-D-glucosidase seems to be a new member of the family 1 of glycosyl hydrolases.
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PMID:Purification, partial amino acid sequence and structure of the product of raucaffricine-O-beta-D-glucosidase from plant cell cultures of Rauwolfia serpentina. 1023 58

Three cellobiases, here called cellobiase A, B, and C, from the culture filtrate of Aspergillus niger A20, were purified by precipitation with ammonium sulphate, gel filtration through Sephadex G-75, and column chromatography of DEAE-cellulose. The purified enzymes were homogeneous on polyacrylamide disk electrophoresis. The mol wt of the purified enzymes were estimated by SDS-gel electrophoresis to be 88,000, 80,000, and 71,000 for cellobiases A, B, and C, respectively. The enzymes were active at pH 4.5 and 55-60 degrees C. The pattern of their amino acid compositions showed high contents of aspartic acid, glutamic acid, threonine, serine, and glycine. The apparent K(m) values for cellobiose were 0.9, 1.63, and 1.0 mM for cellobiases A, B, and C, respectively. Calcium ions stimulated cellobiases B and C, and Co2+ and Mg2+ ions stimulated cellobiase A. The purified enzymes hydrolyzed cellobiose and aryl-beta-D-glucosides, but they had no action on sucrose, maltose, and cellulose. The three cellobiases catalyzed transglycosylase reaction, and the major product formed from cellobiose was tetramer of glucose.
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PMID:Purification and properties of three cellobiases from Aspergillus niger A20. 1032 88

Myrosinase (thioglucoside glucohydrolase; EC 3.2.3.1) is a plant enzyme that hydrolyses glucosinolates, principally to isothiocyanates. Myrosinase was purified to homogeneity in good yield from 8-day-old seedlings of Raphanus sativus (daikon) using a four-step procedure involving chromatographies on anion exchange, hydrophobic Phenyl-Sepharose, gel filtration and concanavalin A-Sepharose. In order to stabilize the enzyme and to avoid excessive peak broadening during chromatography, 30% (v/v) glycerol was added to dialysis and chromatography buffers. The purified enzyme was eluted as a single peak from a gel-filtration sizing column with an apparent molecular mass of 120 kDa. The enzyme was resolved into two subunits with molecular masses of 61 and 62 kDa by SDS/PAGE. Ascorbic acid activated the purified enzyme more than 100-fold. The V(max) and K(m) values for the hydrolysis of allyl glucosinolate (sinigrin) were 2.06 micromol/min per mg of protein and 23 microM in the absence of ascorbate and 280 micromol/min per mg of protein and 250 microM in the presence of 500 microM ascorbate, respectively. As the ascorbate concentration was increased from 50 to 500 microM, the V(max) and K(m) values increased in parallel, and thus the V(max)/K(m) ratio remained constant. Similarly, raising the concentrations of sinigrin increased the concentration of ascorbic acid required for half-maximal activation (K(a)). At a sinigrin concentration of 250 microM, the K(a) for ascorbic acid was 55 microM. Sulphate, a reaction product, was a competitive inhibitor of activity, having a K(i) of 60 mM with respect to sinigrin and of 27 mM with respect to ascorbate. Thus activation of myrosinase from R. sativus by ascorbic acid exemplifies an unusual and possibly unique example of linear 'uncompetitive activation' (i.e. a proportionate increase in V(max) and K(m)) of an enzyme. The enzyme also had beta-glucosidase activity and hydrolysed p-nitrophenyl-beta-d-glucopyranoside.
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PMID:An unusual case of 'uncompetitive activation' by ascorbic acid: purification and kinetic properties of a myrosinase from Raphanus sativus seedlings. 1041 37


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