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:3.2.1.23 (beta-galactosidase)
14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two forms of beta-galactosidase from newborn rat epidermis could be separated by DEAE-cellulose chromatography. Both enzymes showed similar enzymic properties. They had a pH optimum around 3.5--4.5 and the optimal temperature of these enzymes was approximately 60 degrees C. They were not affected by divalent cations, ethylenediaminetetraacetic acid(EDTA) and 2-mercaptoethanol(2-ME), while rho-chloromercuribenzoic acid (PCMB) was a strong inhibitor for each enzyme. These enzymes showed the same Km value (1.25 x 10(-4) M) towards 4-methylumbelliferyl-beta-D-galactoside. However they had different isoelectric points at pH 6.3 and 9.0, respectively. Six different forms of beta-galactosidase activity were found by using isoelectric focusing. When the crude extract was incubated with neuraminidase before electrofocusing, the acidic forms of the enzyme were largely lost and converted to more basic forms without loss of the total activity. This finding suggests the glycoprotein nature of newborn rat epidermal beta-galactosidase.
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PMID:Heterogeneity and some properties of beta-galactosidase from newborn rat epidermis. 11 67

Carboxymethylated beta-galactosidase from Escherichia coli was dissociated at 100 degrees C to form carboxymethylated fragments A and B. The mol.wts. of carboxymethylated fragments A and B were determined by gel filtration to be 64300 and 22400 respectively. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of carboxymethylated fragments A and B that had been pretreated with 2-mercaptoethanol and sodium dodecyl sulphate yielded mol.wts. of 64000 and 22100 respectively. Carboxymethylated fragments A and B had arginine as their C-terminal amino acid. When a crude extract of E. coli M15 was filtered through a column of Sepharose 6B, it was found that carboxymethylated fragment B could restore beta-galactosidase activity when added to fractions having mol.wts. estimated to be 123000, 262000 and 506000. These fractions are referred to as ;complementable fractions'. Similarly, it was found that carboxymethylated fragment A could restore enzyme activity to tractions having mol.wts. estimated to be 63000, 253000 and 506000. Estimates of the molecular weights of the beta-galactosidase activity obtained by restoration with carboxymethylated fragments A and B were made by filtering the active enzyme through another column of Sepharose 6B. The enzyme obtained by complementation with carboxymethylated fragment B, i.e. the complemented enzyme, had mol.wt. 525000, and that obtained with carboxymethylated fragment A had mol.wts. of 525000, 646000 and 2000000. The latter finding suggests that multiple forms of complemented beta-galactosidase can exist.
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PMID:Restoration of beta-galactosidase to Escherichia coli M15. Complementation studies. 41 87

A highly sensitive microassay method and a microscale purification system were developed to isolate the residual acid beta-galactosidase in GM1-gangliosidosis fibroblasts. The sensitivity of the microassay system, composed of a 96-well microplate and a microplate fluorometer, was 100-fold higher than that of the conventional system and the response was linear in the pmole range. Acid beta-galactosidase was characterized as a thiol enzyme which was inactivated by a mercuric compound. This enzyme was completely adsorbed on an Hg-agarose column and was easily eluted from the column by 10 mM 2-mercaptoethanol. The microscale purification system using Con A-Sepharose, PAT-Sepharose, and Hg-agarose column chromatography achieved 565- and 7,970-fold purifications of acid beta-galactosidase with an overall yields of 44% and 45% from normal and GM1-gangliosidosis fibroblasts, respectively. The purified enzyme fractions did not contain any other lysosomal enzyme activities except for a small amount of beta-N-acetylhexosaminidase activity.
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PMID:Acid beta-galactosidase from human fibroblasts. A microscale purification method monitored by a highly sensitive enzyme assay. 308 62

Methods for isolation and purification of beta-galactosidase from Bacillus subtilis, st. IBP-101 are described. The bacterial cells were disrupted by different procedures such as freezing and thawing with subsequent autolysis at 37 degrees C, disrupting in a French press DKM-3 or in ultrasonic disintegrators UZDN-1 (USSR) and Soniprep-150. It is shown that the specific activity and yield of the enzyme depends to a great extent on the disrupting procedure used. The best results were obtained in case of sonication. The preparation was purified by precipitation with ammonium sulphate (25-75% saturated) and chromatography on DEAE-cellulose and DEAE-Sephadex. The purified enzyme had a specific activity of 3155 units per mg protein. The molecular weight of the homogeneous according to gel polyacrylamide electrophoresis preparation was 215,000, as estimated by gel filtration, and 105,000, as estimated by SDS gel electrophoresis. The enzyme retains the activity in the presence of Na+, Mn2+ or Mg2+ ions or the thiolic reagents, dithiothreitol or 2-mercaptoethanol. The pH optimum of the enzyme activity is 6.3 and it is stable in water solutions at pH from 6 to 9 and can be lyophilized. The given preparation of beta-galactosidase has a high affinity for synthetic substrates such as o- and p-nitrophenyl-beta-D-galactopyranosides and 4-methylumbelliferyl-beta-D-galactopyranoside.
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PMID:[Isolation of a highly active preparation of beta-D-galactosidase]. 311 62

A beta-glucosidase (EC 3.2.1.21) was purified to homogeneity from cell-free extracts of an extremely thermophilic anaerobic bacterium. The enzyme has an Mr of 43,000 as determined by molecular-exclusion chromatography, has a pI of 4.55 and shows optimum activity at pH 6.2. The enzyme is active against a wide range of aryl beta-glycosides and beta-linked disaccharides, with beta-galactosidase activity only slightly less than beta-glucosidase activity, and significant beta-xylosidase activity. Lineweaver-Burk plots for p-nitrophenyl beta-glucoside, o-nitrophenyl beta-glucoside and cellobiose substrates are biphasic concave-downwards. Inhibition of the beta-glucosidase by substrates and glucose is negligible. Thermal inactivation follows first-order kinetics, with t1/2 (65 degrees C) 45 h, t1/2 (75 degrees C) 47 min and t1/2 (85 degrees C) 1.4 min and a deactivation energy of 380 kJ/mol at pH 6.2. At pH 7.0, which is the optimum pH for thermostability, t1/2 (75 degrees C) is 130 min. At 75 degrees C, at pH 6.2, the thermostability is enhanced about 8-fold by 10% (w/v) glycerol, about 6-fold by 0.2 M-cellobiose and about 3-fold by 5 mM-dithiothreitol and 5 mM-2-mercaptoethanol.
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PMID:Purification and properties of a stable beta-glucosidase from an extremely thermophilic anaerobic bacterium. 311 33

The isopycnic centrifugation of beta-amylase and the marker enzyme beta-galactosidase was carried out using four salts viz. rubidium chloride, potassium bromide, potassium acetate and lithium bromide. Lithium bromide inactivated both beta-galactosidase and beta-amylase. The high viscosity of potassium acetate gradients necessitated an extremely long centrifugation time. The density profiles obtained with rubidium chloride gradients were sharper and permitted better resolution than potassium bromide gradients. Both enzymes were stable in rubidium chloride gradients, while potassium bromide inactivated beta-galactosidase, even in the presence of 10 mM 2-mercaptoethanol.
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PMID:Advantages of rubidium chloride gradients for isopycnic centrifugation of enzymes. 620 70

A macromolecular aggregate of corticotropin-beta-lipotropin common precursor had been observed in ovine pituitary preparations as an excluded fraction of Sephadex G-200 gel filtration. This fraction could not penetrate a 10% gel during sodium dodecyl sulfate-polyacrylamide gel electrophoresis, when 2-mercaptoethanol or other disulfide-cleaving agents were not present in the buffer used to solubilize the protein preparation prior to the electrophoresis. On a 4.6% gel (acrylamide:bisacrylamide, 20:1), the material migrated as a diffuse band to a position between those of beta-galactosidase (Mr 130 000) and myosin (Mr 200 000). Both observations were consistent with an apparent Mr greatly in excess of that of the corticotropin-beta-lipotropin common precursor reported by many investigators. Neither 5% SDS nor 1% Triton X-100 could dissociate the macromolecular aggregate, but 2-mercaptoethanol and urea, either alone or in combination, were able to dissociate it to two main protein components, one of which was identified as corticotropin-beta-lipotropin with an apparent Mr of 34 000. The fact that urea alone could dissociate this macromolecular aggregate led us to believe that it might be a non-covalent aggregate and that 2-mercaptoethanol probably did not achieve the dissociation through the cleavage of an interchain disulfide bond but by bringing about conformational changes as a result of reduction of intrachain disulfide bonds so that aggregation became unfavorable. Moreover, the dissociation by urea or by 2-mercaptoethanol was found to be irreversible. The origin of the macromolecular aggregate of corticotropin-beta-lipotropin common precursor remains obscure.
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PMID:Characterization of a macromolecular aggregate of ovine pituitary corticotropin-beta-lipotropin common precursor. 626 48

A study of the beta-galactosidase produced by the psychrotrophic bacterium Buttiauxella agrestis has been carried out. This micro-organism was isolated from raw milk and the enzyme isolated using standard methods. Molecular mass was estimated to be 515 kDa. The isoelectric point was close to 4.45. Optimum pH was 7.25. Maximal activity was observed at 50 degrees C and activation energy was estimated to be 39.1 kJ mol-1. Lactose enhanced thermal stability. Using p-nitrophenyl-beta-D-galactopyranoside as the substrate, the Km was 11 mumol l-1 and Vmax was 85 U mg-1 protein. beta-Mercaptoethanol and ethanol were inhibitors; glycerol acted as a complex effector. The enzyme required divalent cations for activity while it was inhibited by EDTA. When the enzyme was immobilized in diethyl aminoethylcellulose the optimum pH of activity was 8. Km was 47 mumol l-1 and Vmax was 96 U mg-1 protein.
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PMID:Isolation and properties of free and immobilized beta-galactosidase from the psychorotrophic enterobacterium Buttiauxella agrestis (strain NC4). 761 19

Nucleophiles activated the catalytic actions of beta-galactosidases with neutral or positively charged substitutions for Glu-461. Aliphatic carboxylic acids increased the rate of hydrolysis of o-nitrophenyl beta-D-galactopyranoside if the pKa values of the carboxyl groups were > approximately 3.5. Amino compounds activated if their pKa values were < approximately 8.5. Imidazole, azide, and 2-mercaptoethanol also activated. Nucleophiles with high pKa values were able to activate the catalysis if the pH was high, and this showed that the lack of activation at pH 7.0 was because of protonation. Kinetic analysis showed that most of the nucleophiles that activated were bound to the active site, since the activation followed Michaelis-Menten type saturation kinetics. The binding seemed to be dependent upon the hydrophobicity; the longer the aliphatic chain, the stronger the binding. Gas-liquid chromatographic analysis showed that adducts of some type were formed during the reactions in the presence of many of the nucleophiles. Three of these adducts were purified and the nucleophiles were found beta-linked to D-galactose. This indicates that if an intermediate covalent bond is formed in the mechanism of beta-galactosidase action and if the nucleophile reacts to displace it, the intermediate covalent bond must have the alpha configuration and involve a group other than Glu-461.
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PMID:beta-Galactosidases of Escherichia coli with substitutions for Glu-461 can be activated by nucleophiles and can form beta-D-galactosyl adducts. 790 53

In the present study, the motilin receptor was characterized by enzymatic digestion studies and by solubilization of the motilin-receptor complex from prelabeled membranes using the anionic detergent cholic acid. Motilin binding was significantly decreased by preincubation of membranes of rabbit antral tissue with trypsin, phospholipase A2, C, D, dithiothreitol and 2-mercaptoethanol but not by neuraminidase and beta-galactosidase. Treatment of prelabeled membranes with 1% cholic acid resulted in solubilization of 24 +/- 5% of the proteins and 65 +/- 3% of the radioactivity. The latter was for 77 +/- 4% due to the presence of the motilin-receptor complex as estimated with PEG-precipitation. Upon gel-filtration on Superose 6 the complex partially dissociated but 43 +/- 3% eluted with macromolecular components in the void volume. This peak was not detected when membranes were first incubated with unlabeled motilin. Further disaggregation was accomplished by the addition of 0.5 M NaCl to the elution buffer. The chromatographic profile then showed a peak of about 370 kDa and a second one of 100 kDa. The latter value probably reflects the molecular mass of a single 125I-motilin-receptor-complex.
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PMID:Solubilization and characterization of motilin-receptor complexes from rabbit antral smooth muscle tissue. 851 70


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