Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Purification of an acid alpha-glucosidase by dextran-gel filtration.

ABSTRACT: 1. The behaviour of rat liver alpha-glucosidases on dextran gel (Sephadex G-100) columns was studied. A ;retardation' of an acid alpha-glucosidase activity was observed. This activity was identified as lysosome alpha-(1-->4)-glucosidase. A single gel-filtration step resulted in a 700-fold purification of the enzyme. The same technique was also used to purify the acid alpha-glucosidase of human kidney. 2. The acid alpha-glucosidases of both tissues show very similar pH optima when tested with maltose or glycogen as substrate.

SUBMITTER: Auricchio F

PROVIDER: S-EPMC1198270 | biostudies-other | 1967 Oct

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Further purification and characterization of the acid alpha-glucosidase.

Project description:1. Centrifugation of rat liver acid glucosidase, which had been purified by adsorption on dextran gel, on a density gradient of sucrose showed the enzyme to be impure. 2. Preliminary purification of the enzyme before the gel filtration improved the final degree of purity of this preparation. Disc gel electrophoresis of this preparation showed a single band of protein. 3. The sedimentation co-efficient and the molecular weight determined on a sucrose gradient were 4.9-5.1s and 76000-83000 respectively for the rat liver enzyme, and 5.6s and 97000 for the acid alpha-glucosidase purified by means of the same procedure from the human kidney. 4. The Michaelis constants of rat liver and human kidney enzyme were 4.7x10(-3)m and 13.6x10(-3)m respectively with maltose as substrate. 5. The enzyme from both tissues was inhibited by tris and by erythritol. The inhibition of the rat liver acid glucosidase by erythritol was competitive.

| S-EPMC1198789 | biostudies-other

Dextran-gel filtration of enzymes in the presence of their high-molecular-weight substrates.

Project description:1. Trypsin and ribonuclease were filtered through dextran gel (Sephadex G-100) columns in the absence and presence of their respective substrates. In the presence of their high-molecular-weight substrates the enzymes emerged earlier from the columns. This appeared to be due to the reversible formation of specific enzyme-substrate complexes. 2. The possibility of separation of an enzyme from other proteins with similar molecular weights was demonstrated with trypsin and cytochrome c in the presence of casein.

| S-EPMC1264828 | biostudies-other

Heterogeneity of pig lysosomal acid alpha-glucosidase. Affinity to Sephacryl S-200 gel and tissue distribution.

Project description:Acid alpha-glucosidase purified from pig liver showed heterogeneity in its affinity to Sephacryl S-200 gel. Acid alpha-glucosidase was separated into two fractions (S1 and S2) by Sephacryl S-200 affinity chromatography. Each fraction contained components at apparent 76 kDa and 67 kDa on SDS/PAGE. The amount of S1 fraction was about 1.3 times that of the S2 fraction. In the kidney the ratio of S1 to S2 fraction was similar to that in the liver. However, the heart contained 1.3 times as much S2 fraction as S1 fraction. The spleen acid alpha-glucosidase consisted mainly of S1 fraction, containing only a 76 kDa component. Immunohistochemically, acid alpha-glucosidase was demonstrated in the macrophages of the spleen. Thus the 76 kDa component in the spleen must come mainly from the macrophages. Lectin-binding analysis was carried out on the components present in the S1 and S2 fractions after electrophoresis and transfer to nitrocellulose sheets. Slight differences in binding observed suggest differences in the structure of the sugar side chains.

| S-EPMC1151505 | biostudies-other

Gel filtration of sialoglycoproteins.

Project description:The role of sialic acid in the gel-filtration behaviour of sialoglycoproteins was investigated by using the separated isoenzymes of purified human liver alpha-L-fucosidase and several other well-known sialic acid-containing glycoproteins (fetuin, alpha1-acid glycoprotein, thyroglobulin and bovine submaxillary mucin). For each glycoprotein studied, gel filtration of its desialylated derivative gave an apparent molecular weights much less than that expected just from removal of sialic acid. For the lower-molecular-weight glycoproteins (fetuin and alpha1-acid glyocprotein), gel filtration of the sialylated molecules led to apparent molecular weights much larger than the known values. The data indicate that gel filtration cannot be used for accurately determining the molecular weights of at least some sialoglycoproteins.

| S-EPMC1185776 | biostudies-other

Isolation of acid tolerant lactic acid bacteria and evaluation of α-glucosidase inhibitory activity.

Project description:In this study, lactic acid bacteria strains (LABs) were isolated from Korean traditional fermented food and examined as potential probiotics using in vitro methods. Ten LAB strains survived in de Man, Rogosa and Sharpe broth adjusted to pH 2.5 were tested for resistance to acidic conditions and bile, antimicrobial activity, and α-glucosidase inhibitory activity. Among them, strain MBEL1397 showed antimicrobial activity against Bacillus cereus and exhibited survival rates of over 97% in acidic and bile conditions. The α-glucosidase inhibitory activity was 3.91 ± 0.25%, corresponding to approximately 2.3 times higher than that of acarbose. MBEL1397 was susceptible to ampicillin, erythromycin, and penicillin G and identified as Lactobacillus sakei. It was deposited to Korean Collection for Type Culture (KCTC) as KCTC14037BP. In conclusion, these results demonstrate that L. sakei MBEL1397 might be prominent probiotics with potential hypoglycemic effects.

| S-EPMC7347726 | biostudies-literature

Chromatography of transforming deoxyribonucleic acid on methylated albumin and by gel filtration.

Project description:1. Native DNA from two strains of Bacillus subtilis was chromatographed by stepwise elution from MAK (methylated albumin on kieselguhr). 2. Transforming activity was confined to two out of the three main fractions, activity being distributed between the two peaks differently for DNA from the different strains. 3. Fractionation of DNA from both strains on 2% agarose gel gave two components. Approx. 75% of the material was eluted within the void volume of the column. Approx. 25% of the material consisted of degradation products of lower molecular weight. 4. Chromatography on MAK of the material of high molecular weight eluted from agarose gel gave a number of peaks differing in molecular weight, indicating that degradation of the DNA takes place during chromatography on MAK. 5. The distribution of transforming activity among the fractions from MAK suggests that degradation occurs preferentially in certain regions of the DNA.

| S-EPMC1166155 | biostudies-other

Rescue of advanced Pompe disease in mice with hepatic expression of secretable acid α-glucosidase

Project description:Pompe disease is a neuromuscular disorder caused by mutations in the gene encoding for the lysosomal enzyme acid α-glucosidase (GAA). GAA converts lysosomal glycogen to glucose, and its deficiency leads to pathologic glycogen accumulation. Enzyme replacement therapy (ERT) is the only available treatment for Pompe disease at the moment with several shortcomings. We have shown that liver expression of secGAA has better therapeutic efficacy than non-engineered GAA after long-term treatment of four months old Gaa-/- mice with low vector doses. Based on those results, we have treated severely affected nine months old Gaa-/- mice with the AAV-secGAA vector and followed the animals for nine months thereafter. At the end of the study, AAV-treated Gaa-/- mice showed complete rescue of the Pompe phenotype. Transcriptomic profiling of skeletal muscle highlighted mitochondrial bioenergetics defects, supported by electron microscopy, western blotting and biochemical findings, which were partially corrected after AAV treatment. Together, these results provide insight into the reversibility of advanced Pompe disease in the Gaa-/- mouse model via liver gene transfer of secGAA.

2021-04-26 | GSE150935 | GEO

Heat-stable protein that stimulates acid alpha-glucosidase.

Project description:A hot-water extract of bovine spleen and guinea pig liver exhibited the ability to enhance acid alpha-glucosidase activity, with methylumbelliferyl alpha-glucoside, glycogen or maltose as substrate. The level of activator required for maximal stabilization was similar for all three substrates, indicating direct action on the enzyme rather than on substrate. The stimulator was partially purified by chromatography with gel-permeation (apparent Mr 20,000-24,000), ion-exchange and C4 reverse-phase columns. It was retained by a narrow-pore dialysis tubing and destroyed by treatment with Pronase, and is presumably a protein. The stimulating protein protected the enzyme against denaturation by heat or incubation with a buffer of high ionic strength in the absence of substrate. RNA inhibited the enzyme, and the activator protein was able to counteract the effect. Activating material was found in a variety of mouse and rat tissues, as well as human urine.

| S-EPMC1133662 | biostudies-other

Crystallization and preliminary X-ray analysis of Streptococcus mutans dextran glucosidase.

Project description:Dextran glucosidase from Streptococcus mutans is an exo-hydrolase that acts on the nonreducing terminal alpha-1,6-glucosidic linkage of oligosaccharides and dextran with a high degree of transglucosylation. Based on amino-acid sequence similarity, this enzyme is classified into glycoside hydrolase family 13. Recombinant dextran glucosidase was purified and crystallized by the hanging-drop vapour-diffusion technique using polyethylene glycol 6000 as a precipitant. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 72.72, b = 86.47, c = 104.30 A. A native data set was collected to 2.2 A resolution from a single crystal.

| S-EPMC2376310 | biostudies-literature