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)

beta-D-Glucopyranosyl-(1S and 1R)-epoxyethanes (I and II), 1-(beta-D-glucopyranosyl)-(2R and 2S)-2,3-epoxypropanes (III and IV), beta-D-glucopyranosyl isothiocyanate (V) and beta-D-galactopyranosylepoxyethane (VI) are active-site-directed irreversible inhibitors of sweet-almond beta-glucosidase B (beta-D-Glucoside glucohydrolase, EC 3.2.1.21). Formation of the covalent bond is preceded by the binding of these inhibitors in the active site of the enzyme. This is testitified by the competitive character of inhibition of beta-glucosidase component B by compounds I-VI at the early period and by the protection of the enzyme from inactivation by its competitive inhibitors D-glucose and 1,5-D-gluconolactone. Epoxides I-IV are bound covalently with componet B at a molar ratio 1 : 1 as shown with the aid of 14C-labelled inhibitors. The release of the label from modified enzyme (E-I covalent) by treatment with hydroxylamine suggests the formation of an ester bond between inhibitors I-IV and the carboxyl group of the enzyme active site. The pH dependence curve of the inactivation rate of beta-glucosidase B is of a bell-shaped form for V and of a sigmoid character for I-IV and points to the involvement of the active site groups with pKa 5.6-5.9 and 4.2-4.4.
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PMID:Specfic irreversible inhibition of sweet-almond beta-glucosidase by some beta-glycopyranosylepoxyalkanes and beta-d-glucopyranosyl isothiocyanate. 0 36

1. Whereas only beta-glucosidase A (beta-D-glucoside glucohydrolase, EC 3.2.1.21) was produced by the tropical fungus Botryodiplodia theobromae Pat. (I.M.I. 115626; A.T.C.C. 26123) in young cultures containing D-cellobiose as carbon source, lower-Mr forms (B, C and D) were found in older cultures when the pH had drifted from the initial value of pH 6.2 to pH 7.9. 2. The Michaelis constants (Km) of the various molecular forms of the enzyme were 0.30 +/- 0.03 mM-, 0.26 +/- 0.01 mM-, 0.20 +/- 0.02 mM- and 0.16 +/- 0.01 mM-o-nitrophenyl beta-D-glucopyranoside for beta-glucosidase forms A (Mr 320,000), B (Mr 160,000), C (Mr 80,000) and D (Mr 40,000) respectively. 3. Only beta-glucosidase D showed substrate inhibition. 4. Only L-arginine was found as the N-terminal residue, and beta-glucosidase A contained 31.7 +/- 0.6 mol of N-terminal L-arginine/mol of the enzyme. 5. Storage of purified beta-glucosidase A under mildly alkaline conditions caused its dissociation into the lower-Mr forms, whereas adjustment of the pH of a solution of beta-glucosidase A to pH 12.0 with 1 M-NaOH led to complete inactivation on incubation at 40 degrees C for 1 h and to the release of 25.2 +/- 1.5 mol of inorganic phosphate/mol of the enzyme. 6. O-Phospho-L-serine was isolated from the acid-hydrolysis product of beta-glucosidase A but not from that of beta-glucosidase D. 7. Reduction and carboxamidomethylation of the various forms of beta-glucosidase gave only one enzymically inactive protein with an Mr of 10,000-11,000. 8. After partial succinylation (3-carboxypropionylation) of beta-glucosidase D at pH 5.0 and removal of the precipitated protein formed, the supernatant solution contained beta-glucosidase components similar to the other molecular forms (A, B and C) and an aggregate (beta-glucosidase Xs) that gave a positive result in the alkaline hydroxylamine test, whereas N-succinylated beta-glucosidase D, an aggregate (form Xp) that behaved like beta-glucosidase Xs and traces of forms A, B and C were found by gel filtration of the solution of the precipitate solubilized at neutral pH (7.0-7.7). 9. These observations are discussed in terms of the proposed octameric structure of beta-glucosidase A based on the result of electron microscopy [Umezurike (1975) Biochem. J. 145, 361-368].
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PMID:The octameric structure of beta-glucosidase from Botryodiplodia theobromae Pat. 190 26

The bglA gene from Bacillus polymyxa encodes a beta-glucosidase able to hydrolyze p-nitrophenyl-beta-D-glucopyranoside (PNPG), and 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-gal), chromogenic substrates for beta-glucosidases and beta-galactosidases respectively. A plasmid carrying the blgA gene inserted in vector pUC18 was mutagenized in vitro with hydroxylamine, and subsequently used to transform E. coli selecting for the ampicillin resistance conferred by the cloning vector. The transformants were screened on petri dishes for mutations causing inability to hydrolyze either one of the two substrates, and for mutations increasing resistance of the enzyme to thermal inactivation. The isolation of several mutants with such characteristics suggests that the simple procedure used here can be applied to generate modifications of enzymatic properties that fit specific industrial requirements.
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PMID:Random mutagenesis of a plasmid-borne glycosidase gene and phenotypic selection of mutants in Escherichia coli. 767 73

The broad-specificity cyanogenic beta-D-glucosidase (beta-D-glucoside glucohydrolase, EC 3.2.1.21) (linamarase) from Manihot esculenta Crantz (cassava) was kinetically characterized in mixed substrate systems and with the transition-state analogue glucono(1-5)lactone and a series of 1-thio substrate analogues. The results indicate a common catalytic and a common sugar binding site in the enzyme for all of the investigated substrates. Kinetic parameters of the hydrolysis of linamarin and p-nitrophenyl beta-D-glucopyranoside were determined over the pH range 3.5-9.0. The pH-dependence curves gave apparent pK values of 4.5 (4.6) and 7.1 (7.3) for the free enzyme, while values of 4.5 (3.7) and 9.3 were obtained for the enzyme-substrate complexes, using either linamarin or p-nitrophenyl beta-D-glucopyranoside as the substrate. Kinetic analysis of the modification indicated that one molecule of water-soluble carbodiimide or Woodward's reagent K is required to bind to the enzyme for inactivation. The enzyme was protected against inactivation by the competitive inhibitors p-nitrothiophenyl beta-D-glucopyranoside, beta-D-glucopyranosylamine, and glucono(1-5)lactone. Spectrophotometric analysis at 340 nm showed that from the three carboxylate groups modified by Woodward's reagent K essentially one was protected by p-nitrothiophenyl beta-D-glucopyranoside. During modification Vmax decreased to 30% of that of the unmodified enzyme and Km remained unchanged. The pH dependence of inactivation showed the involvement of a group with a pK value of 4.6, indicating the modification of a carboxyl residue essential for activity. Treatment of the enzyme with the histidine-group-specific reagent diethylpyrocarbonate resulted in 80% loss of enzyme activity, in biphasic kinetics. A treatment with 0.5 M hydroxylamine at pH 7.0 regenerated 92% of the original enzyme activity. The presence of the competitive inhibitor beta-D-glucopyranosylamine protected the enzyme against inactivation, preventing the modification of one histidine residue. Statistical analysis of the residual fractional activity against the number of modified residues indicated that the modification of one histidine is responsible for 40-50% of the inactivation. The pH dependence of the inactivation gave a pK value of 7.0 for the histidine group upon which the activity depends. During modification, Vmax decreased to 30% and Km decreased to 50% of the original values.
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PMID:Investigation of the active site of the cyanogenic beta-D-glucosidase (linamarase) from Manihot esculenta Crantz (cassava). I. Evidence for an essential carboxylate and a reactive histidine residue in a single catalytic center. 794 86

The inhibition of beta-glucosidase from Trichoderma reesei QM 9414 by several specific reagents was studied. Diethylpyrocarbonate (DEP) nearly abolished the enzyme activity at concentrations above 10 mM. The presence of substrate or analogs protected the enzyme against inactivation. The reaction followed pseudo-first order kinetics with a second-order rate constant of 0.02 mM-1.min-1. The pH-dependence of the inactivation showed the involvement of a group with a pK of 5.2. Difference spectra at 242 nm and the reversal of the inactivation in the presence of 1 M hydroxylamine indicated the modification of histidine residues. Statistical analysis of residual fractional activity versus the number of modified histidine residues indicated that one histidine residue is essential for catalysis. p-Hydroxymercuribenzoate completely inhibited the enzyme at concentrations of the reagent above 2 mM. Substrate or analogs protected the enzyme against inactivation. The reaction followed pseudo-first order kinetics with a second-order rate constant of 0.002 mM-1.min-1. Treatment of the modified enzyme with 5,5'-dithio-bis(2-nitrobenzoic acid) (DTNB) showed that one cysteine residue was essential for activity. At pH 5.0 2-ethoxy-1-ethoxy-carbonyl-1,2-dihydroquinoline (EEDQ) inactivated the enzyme according to pseudo-first order kinetics with a second-order rate constant of 0.12 min-1. The pH-dependence of the inactivation showed the involvement of a group with a pK of 5.64, indicating the modification of a carboxyl group essential for activity.
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PMID:Chemical modification of beta-glucosidase from Trichoderma reesei QM 9414. 811 32

Mutations enhancing the thermostability of beta-glucosidase A of Bacillus polymyxa, a family 1 glycosyl hydrolase, have been obtained after hydroxylamine mutagenesis of a plasmid containing the bglA gene, transformation of Escherichia coli with the mutagenized plasmid, and identification of transformant colonies that showed beta-glucosidase activity after a thermal treatment that inactivated the wild-type enzyme. Two additive mutations have been characterized that cause replacement of glutamate at position 96 by lysine and of methionine at position 416 by isoleucine respectively. The thermoresistant mutant enzymes showed increased resistance to other denaturing agents, such as pH and urea, while their kinetic parameters did not change. CD spectra indicated that the E96K replacement caused an increase in alpha-helix content. The observed effect of the M416I mutation is consistent with the lower content of cysteine and methionine found in family 1 enzymes of thermophilic species compared with similar ones from mesophilic organisms.
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PMID:Amino acid substitutions enhancing thermostability of Bacillus polymyxa beta-glucosidase A. 861 77

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