Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

The separation and characterization of the methylumbelliferyl beta-galactosidases of human liver.

ABSTRACT: 1. A previously uncharacterized form of human liver acid beta-galactosidase (EC 3.2.1.23), possibly a dimer of molecular weight 160 000, was resolved by gel filtration. It has the same ability to hydrolyse GM1 ganglioside as the two other acid beta-galactosidase forms. 2. The low-molecular-weight forms of acid beta-galactosidase undergo salt-dependent aggregation. 3. The high-molecular-weight component may consist of the low-molecular-weight forms bound to membrane fragments. It can be converted completely into a mixture of these forms. 4. The neutral beta-galactosidase activity can be resolved into two forms by DEAE-cellulose chromatography. They differ in their response to Cl-ions. 5. A new nomenclature is suggested for the six beta-galactosidases so far found in human liver. 6. The enzymic constituents of the beta-galactosidase bands resolved by electrophoresis were re-examined. The A band contains three components. A two-dimensional electrophoretic procedure for resolving the A band is described. 7. The effect of neuraminidase treatment on the behaviour of beta-galactosidases in various separation systems is examined.

SUBMITTER: Cheetham PS

PROVIDER: S-EPMC1163830 | biostudies-other | 1976 Jul

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Human small-intestinal beta-galactosidases. Separation and characterization of one lactase and one hetero beta-galactosidase.

Project description:1. Two beta-galactosidases from human small-intestinal mucosa were separated by gel-filtration chromatography and the properties of the two enzymes were studied. Lactose and four hetero beta-galactosides were used as substrates. 2. One of the enzymes was particle-bound and could be partially solubilized with papain. Of the substrates hydrolysed by this enzyme, lactose was hydrolysed most rapidly. This enzyme is thus essentially a disaccharidase and is named lactase. It is presumably identical with the ;lactase 1' described earlier. 3. The other enzyme was mainly soluble and hydrolysed all artificial substrates used, whereas no lactase activity could be detected. This enzyme has therefore been designated hetero beta-galactosidase. 4. p-Chloromercuribenzoate (0.1mm) inhibited the hetero beta-galactosidase completely but did not influence the activity of the lactase. Tris was a competitive inhibitor of both enzymes. 5. The residual lactase activity in the mucosa of lactose-intolerant patients may be exerted by a small amount of remaining lactase as such, or possibly by a third enzyme with a more acid pH optimum.

| S-EPMC1184862 | biostudies-other

Characterization of purified human liver acid beta-D-galactosidases A2 and A3.

Project description:1. Human liver acid beta-galactosidase A2 and A3 were isolated by chromatography on concanavalin A-Sepharose 4B, Sepharose 6B, and Sepharose 4B-6-aminohexyl 1-thio-beta-D-galactopyranoside. beta-Galactosidase A2 and A3 were purified to final specific activities of 45.5 and 20.6 mumol/min per mg respectively with 4-methylumbelliferyl beta-D-galactopyranoside as substrate. 2. Form A2 had a mol.wt. of 150000 +/- 15000 (gel filtration) and appeared as a single band of protein (mol.wt 65000 +/- 1000) on electrophoresis in the presence of sodium dodecyl sulphate. 3. Form A3 had a mol.wt. (gel filtration) of 660000 +/- 66000. On electrophoresis in the presence of sodium dodecyl sulphate, form A3 appeared as a major band of protein (72% of total) of mol.wt. 65000 +/- 1000 and minor protein bands of mol.wt. 44000 +/- 1000 and 26,000 +/- 1000 and 22000 +/- 1000. 4. Gel-filtration chromatography of purified beta-galactosidase A3 generated approximately equal amounts of forms A3 and A2. beta-Galactosidase A1 was not detected by gel-filtration chromatography of partially or highly purified preparations of forms A2 and A3. 5. Both forms A2 and A3 had identical isoelectric points of 4.42 +/- 0.02. The data suggest that forms A2 and A3 are dimeric and multimeric forms of beta-galactosidase A1. 6. Amino acid analysis of beta-galactosidase A2 gave a ratio of acidic to basic amino acids of 2.6:1. 7. beta-Galactosidase A2 contained 7.5% carbohydrate by weight and sialic acid, D-galactose, D-glucosamine and D-mannose were present in the molar proportions 1.1:1.0:1.7:2.7.

| S-EPMC1186052 | biostudies-other

Characterization of GH2 and GH42 β-galactosidases derived from bifidobacterial infant isolates.

Project description:Bifidobacteria are among the first and most abundant bacterial colonizers of the gastrointestinal tract of (breast-fed) healthy infants. Their success of colonising the infant gut is believed to be, at least partly, due to their ability to metabolize available carbon sources by means of secreted or intracellular glycosyl hydrolases (GHs). Among these, β-galactosidases are particularly relevant as they allow bifidobacteria to grow on β-galactosyl-linked saccharidic substrates, which are present in copious amounts in the milk-based diet of their infant host (e.g. lactose and human milk oligosaccharides). In the present study we employed an in silico analysis to identify GH family 2 and 42 β-galactosidases encoded by typical infant-associated bifidobacteria. Comparative genome analysis followed by characterisation of selected β-galactosidases revealed how these GH2 and GH42 members are distributed among these infant-associated bifidobacteria, while their hydrolytic activity towards growth substrates commonly available in the infant gut were also assessed.

| S-EPMC6339635 | biostudies-literature

β-Galactosidases from a Sequence-Based Metagenome: Cloning, Expression, Purification and Characterization.

Project description:Stabilization ponds are a common treatment technology for wastewater generated by dairy industries. Large proportions of cheese whey are thrown into these ponds, creating an environmental problem because of the large volume produced and the high biological and chemical oxygen demands. Due to its composition, mainly lactose and proteins, it can be considered as a raw material for value-added products, through physicochemical or enzymatic treatments. β-Galactosidases (EC 3.2.1.23) are lactose modifying enzymes that can transform lactose in free monomers, glucose and galactose, or galactooligosacharides. Here, the identification of novel genes encoding β-galactosidases, identified via whole-genome shotgun sequencing of the metagenome of dairy industries stabilization ponds is reported. The genes were selected based on the conservation of catalytic domains, comparing against the CAZy database, and focusing on families with β-galactosidases activity (GH1, GH2 and GH42). A total of 394 candidate genes were found, all belonging to bacterial species. From these candidates, 12 were selected to be cloned and expressed. A total of six enzymes were expressed, and five cleaved efficiently ortho-nitrophenyl-β-galactoside and lactose. The activity levels of one of these novel β-galactosidase was higher than other enzymes reported from functional metagenomics screening and higher than the only enzyme reported from sequence-based metagenomics. A group of novel mesophilic β-galactosidases from diary stabilization ponds' metagenomes was successfully identified, cloned and expressed. These novel enzymes provide alternatives for the production of value-added products from dairy industries' by-products.

| S-EPMC7823827 | biostudies-literature

Human small-intestinal -galactosidases. Separation and characterization of three forms of an acid -galactosidase.

Project description:1. An acid beta-galactosidase, optimum pH4.0-4.5, in the human small-intestinal mucosa was separated and characterized. 2. Autolysis of mucosal homogenates at acid pH inactivated the lactase and hetero beta-galactosidase; the total activity of the acid beta-galactosidase was only slightly depleted, but a greater proportion of the enzyme was solubilized by this treatment. 3. Separation on a Sephadex G-200 column revealed that the acid beta-galactosidase could occur in at least three different forms, probably representing monomer, dimer and octamer or polymer of the enzyme. 4. The properties of the different forms of the acid beta-galactosidase were studied with regard to pH optimum, K(m), rate of hydrolysis of different substrates, and sensitivity to p-chloromercuribenzoate and tris as inhibitors. All these properties were the same for the different forms of the enzyme. 5. The acid beta-galactosidase hydrolyses lactose as well as hetero beta-galactosides and contributes to the lactase activity of intestinal biopsies also when measured at pH 6. This enzyme may therefore be responsible for a considerable part of the residual lactase activity found in lactose-intolerant patients.

| S-EPMC1176570 | biostudies-other

Enzymes of Trichomonas foetus. Separation and properties of two beta-galactosidases.

Project description:The beta-galactosidase activity in extracts of Trichomonas foetus is separable into two fractions by gel filtration on Sephadex G-200. When o-nitrophenyl beta-d-galactoside is used as substrate the first fraction to be eluted, beta-galactosidase 1, has 50 times the activity (units per mg of protein) of the crude preparation. This fraction is activated by Mn(2+) and Co(2+) and inhibited by Hg(2+) and EDTA. In the presence of Mn(2+) the pH optimum for the hydrolysis of o-nitrophenyl beta-d-galactoside or lactose is 5.8-6.0. beta-Galactosidase 1 is an exoglycosidase that releases beta-linked galactose joined to aliphatic and various carbohydrate aglycones. Hydrolysis is prevented, however, by a substituent on either the subterminal sugar or the terminal non-reducing beta-galactosyl residue in an oligosaccharide. The second fraction, beta-galactosidase 2, is not activated by metal ions or inhibited by EDTA and has a broad pH optimum from 4.5 to 6.0.

| S-EPMC1179017 | biostudies-other

Rat small-intestinal beta-galactosidases. Separation by ion-exchange chromatography and gel filtration.

Project description:1. The chromatography of rat small-intestinal beta-galactosidase activities on gel-filtration and ion-exchange columns has been studied. Five different substrates were used to measure beta-galactosidase activity (lactose, phenyl beta-galactoside, o-nitrophenyl beta-galactoside, p-nitrophenyl beta-galactoside and 6-bromo-2-naphthyl beta-galactoside) and the activity was measured at one acid and one more neutral pH value. 2. By gel filtration one acid beta-galactosidase, hydrolysing lactose and the hetero-beta-galactosides at about the same rate, and one more neutral beta-galactosidase, hydrolysing lactose much more rapidly than the hetero-beta-galactosides, were separated. 3. By ion-exchange chromatography the acid enzyme was fractionated into two components. These may be individual enzymes or different forms of the same enzyme.

| S-EPMC1198588 | biostudies-other

Two β-galactosidases from the human isolate Bifidobacterium breve DSM 20213: molecular cloning and expression, biochemical characterization and synthesis of galacto-oligosaccharides.

Project description:Two β-galactosidases, β-gal I and β-gal II, from Bifidobacterium breve DSM 20213, which was isolated from the intestine of an infant, were overexpressed in Escherichia coli with co-expression of the chaperones GroEL/GroES, purified to electrophoretic homogeneity and biochemically characterized. Both β-gal I and β-gal II belong to glycoside hydrolase family 2 and are homodimers with native molecular masses of 220 and 211 kDa, respectively. The optimum pH and temperature for hydrolysis of the two substrates o-nitrophenyl-β-D-galactopyranoside (oNPG) and lactose were determined at pH 7.0 and 50°C for β-gal I, and at pH 6.5 and 55°C for β-gal II, respectively. The kcat/Km values for oNPG and lactose hydrolysis are 722 and 7.4 mM-1s-1 for β-gal I, and 543 and 25 mM-1s-1 for β-gal II. Both β-gal I and β-gal II are only moderately inhibited by their reaction products D-galactose and D-glucose. Both enzymes were found to be very well suited for the production of galacto-oligosaccharides with total GOS yields of 33% and 44% of total sugars obtained with β-gal I and β-gal II, respectively. The predominant transgalactosylation products are β-D-Galp-(1→6)-D-Glc (allolactose) and β-D-Galp-(1→3)-D-Lac, accounting together for more than 75% and 65% of the GOS formed by transgalactosylation by β-gal I and β-gal II, respectively, indicating that both enzymes have a propensity to synthesize β-(1→6) and β-(1→3)-linked GOS. The resulting GOS mixtures contained relatively high fractions of allolactose, which results from the fact that glucose is a far better acceptor for galactosyl transfer than galactose and lactose, and intramolecular transgalactosylation contributes significantly to the formation of this disaccharide.

| S-EPMC4121272 | biostudies-literature

Purified human liver acid beta-D-galactosidases possessing activity towards G(M1)-ganglioside and lactosylceramide.

Project description:Our studies with purified human liver acid beta-D-galactosidases (EC 3.2.1.23) indicate that 4-methylumbelliferyl beta-D-galactosidase and G(M1)-ganglioside beta-D-galactosidase activities are identical with lactosylceramidase II activity. Evidence for this includes co-purification of all enzyme activities by affinity chromatography to yield a single band on polyacrylamide-gel electrophoresis and coincident elution from Sepharose 6B of all three enzyme activities.

| S-EPMC1164944 | biostudies-other

Heterologous expression and characterization of an Arabidopsis β-l-arabinopyranosidase and α-d-galactosidases acting on β-l-arabinopyranosyl residues.

Project description:The major plant sugar l-arabinose (l-Ara) has two different ring forms, l-arabinofuranose (l-Araf) and l-arabinopyranose (l-Arap). Although l-Ara mainly appears in the form of α-l-Araf residues in cell wall components, such as pectic α-1,3:1,5-arabinan, arabinoxylan, and arabinogalactan-proteins (AGPs), lesser amounts of it can also be found as β-l-Arap residues of AGPs. Even though AGPs are known to be rapidly metabolized, the enzymes acting on the β-l-Arap residues remain to be identified. In the present study, four enzymes, which we call β-l-ARAPASE (APSE) and α-GALACTOSIDASE 1 (AGAL1), AGAL2, and AGAL3, are identified as those enzymes that are likely to be responsible for the hydrolysis of the β-l-Arap residues in Arabidopsis thaliana. An Arabidopsis apse-1 mutant showed significant reduction in β-l-arabinopyranosidase activity, and an apse-1 agal3-1 double-mutant exhibited even less activity. The apse-1 and the double-mutants both had more β-l-Arap residues in the cell walls than wild-type plants. Recombinant APSE expressed in the yeast Pichia pastoris specifically hydrolyzed β-l-Arap residues and released l-Ara from gum arabic and larch arabinogalactan. The recombinant AGAL3 also showed weak β-l-arabinopyranosidase activity beside its strong α-galactosidase activity. It appears that the β-l-Arap residues of AGPs are hydrolysed mainly by APSE and partially by AGALs in Arabidopsis.

| S-EPMC5853685 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data