Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “GALACTOSIDASE”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,333 records · Page 74Linked to original sources
PubMed2000

Appearance of biomarkers of in vitro ageing after successive stimulation of WI-38 fibroblasts with IL-1alpha and TNF-alpha: senescence associated beta-galactosidase activity and morphotype transition.

Sublethal oxidative stresses increase the proportions of human fibroblasts positive for senescence associated beta-galactosidase activity and accelerate the transition in the fibroblast morphotypes characterising fibroblast ageing. Stimulation of fibroblasts with TNF-alpha or IL-1alpha transiently increases the production of reactive oxygen species (ROS) in human fibroblasts. Here we propose that repeated stimulation of WI-38 fibroblasts with TNF-alpha or IL-1alpha can generate enough ROS to accelerate the transition in the fibroblast morphotypes and increase the proportion of cells positive for senescence associated beta-galactosidase activity. The involvement of ROS is suggested by experiments where the stimulation of fibroblasts with TNF-alpha or IL-1alpha are performed in the presence of N-acetylcysteine which increases the intracellular antioxidant potential. It is proposed that the decrease in the proportions of morphotypes I and II, and the increase in the proportions of morphotypes III to VI observed after successive stimulation with TNF-alpha or IL1-alpha is attributed to an increased ROS production occurring during the stimulation.

Acetylcysteine↗
PubMed2001

Local and global cerebral blood flow and glucose utilization in the alpha-galactosidase A knockout mouse model of Fabry disease.

Fabry disease is an X-linked lysosomal disorder characterized by deficient alpha-galactosidase A activity and intracellular accumulations of glycosphingolipids, mainly globotriaosylceramide (Gb3). Clinically, patients occasionally present CNS dysfunction. To examine the pathophysiology underlying brain dysfunction, we examined glucose utilization (CMR(glc)) and cerebral blood flow (CBF) globally and locally in 18 brain structures in the alpha-galactosidase A gene knockout mouse. Global CMR(glc) was statistically significantly reduced by 22% in Fabry mice (p < 0.01). All 18 structures showed decreases in local CMR(glc) ranging from 14% to 33%. The decreases in all structures of the diencephalon, caudate-putamen, brain stem, and cerebellar cortex were statistically significant (p < 0.05). Global cerebral blood flow (CBF) and local CBF measured in the same 18 structures were lower in Fabry mice than in control mice, but none statistically significantly. Histological examination of brain revealed no cerebral infarcts but abundant Gb3 deposits in the walls of the cerebral vessels with neuronal deposits localized to the medulla oblongata. These results indicate an impairment in cerebral energy metabolism in the Fabry mice, but one not necessarily due to circulatory insufficiency.

Animals↗
PubMed2000

Alizarin-beta-D-galactoside: a new substrate for the detection of bacterial beta-galactosidase.

We describe the synthesis of a new substrate for the detection of bacterial beta-galactosidase. This substrate, alizarin-beta-D-galactoside, is readily hydrolysed to release alizarin which complexes with various metal ions to form brightly coloured chelates. A total of 367 strains of Gram-negative bacteria were examined for their ability to hydrolyse three chromogenic substrates: alizarin-beta-D-galactoside (Aliz-gal), cyclohexenoesculetin-beta-D-galactoside (CHE-gal) and 5-bromo-4-chloro-3-indolyl-beta-D-galactoside (X-gal). A total of 182 strains (49.6%) were found to hydrolyse at least one of the three substrates. All of these 182 strains (100%) hydrolysed Aliz-gal whereas only 170 (93.4%) and 173 (95.1%) hydrolysed CHE-gal and X-gal, respectively. We conclude that alizarin-beta-D-galactoside is a highly sensitive substrate for the demonstration of beta-galactosidase.

Anthraquinones↗
PubMed2002

Polarity of alpha-galactosidase A uptake by renal tubule cells.

BACKGROUND: Congenital absence of alpha-galactosidase in Fabry disease leads eventually to renal failure. Fabry disease is an attractive candidate for gene therapy, but uptake mechanisms of the enzyme must be understood for it to be used in treating patients with Fabry disease. METHODS: Immortalized human renal epithelial cells from three regions of the tubule were grown in culture on collagen-coated Transwell filters and were incubated with recombinant alpha-galactosidase protein placed at either the luminal or basolateral side of the cells. Uptake into cells was measured, and kinetic studies were performed. Blocking experiments were done with mannose 6-phosphate. RESULTS: Uptake from the basolateral side of the filters predominated in all three cell types. Only in distal tubule cells was mannose 6-phosphate able to block uptake to any degree. The kinetic data reveal a high Km for both luminal and basolateral cell surfaces. CONCLUSIONS: These data suggest that to correct the renal phenotype in Fabry disease, high levels of the enzyme will be need to be delivered to kidney cells. This will likely best be achieved with local administration of a vector containing the transgene directly to the kidney.

Cell Polarity↗
PubMed1988

Galactosylceramide-beta-galactosidase deficiency in association with cherry red spot.

A 13-month-old white girl was the product of a normal pregnancy and delivered by caesarean section for breech presentation. Regression of motor milestones started by 11 months, when delayed language development was also noted. She was normocephalic without major dysmorphic features or organomegaly. Fundus examination disclosed a subtle cherry red spot bilaterally. No startle response was elicited. By 17 months she was extremely irritable and unable to tolerate liquids; there was symmetrical spasticity and florid cherry red spots. She died at 18 months of age. A systematic search for conditions associated with a cherry red spot was unrevealing. The absence of galactosylceramide galactosidase activity was unexpected and was confirmed on three occasions in two laboratories. Lactosylceramide I content, an enzyme thought to be identical to galactosylceramide-beta-galactosidase, was significantly decreased. The presence of a cherry red spot in Krabbe's disease, indicative of neuronal storage, has not been previously recognized. The existence of this variant has implications for genetic and biochemical studies.

Brain↗
PubMed1983

Intestinal glycosidase activities in one adult and two suckling echidnas: absence of a neutral lactase (beta-D-galactosidase).

The activities of various glycosidases in homogenates of the small-intestinal mucosa of one adult and two suckling echidnas, Tachyglossus aculeatus, were investigated. The activities of lactase (beta-D-galactosidase), beta-N-acetylglucosaminidase, neuraminidase and alpha-L-fucosidase were higher in the sucklings than in the adult animal. Maltase and isomaltase activities were lower. Sucrase and cellobiase activities were absent or present in trace amounts only. The lactase activity had a pH optimum of 4.0-4.5, was predominantly in the soluble fraction following ultracentrifugation and was inhibited by p-chloromercuribenzene sulfonate, suggesting that it was due to a lysosomal acid beta-galactosidase and not a brush-border neutral lactase. The maltase activity of the sucklings also had the characteristics predominantly of a lysosomal acid hydrolase. It is proposed that in suckling echidnas, the oligosaccharides (mainly neuraminyllactose and fucosyllactose) of the mother's milk are digested intracellularly by lysosomal enzymes, rather than at the brush border, of the epithelial cells of the small-intestinal mucosa.

Animals↗
PubMed1970

Globoid cell leucodystrophy (Krabbe's disease): deficiency of galactocerebroside beta-galactosidase.

Profound deficiency of a specific enzyme, galactocerebroside beta-galactosidase, has been demonstrated in the brains, liver, and spleen of three patients with Krabbe's globoid cell leucodystrophy. The activity of this enzyme was normal in a variety of other cerebral diseases, including those with similarly devasted white matter. The lack of enzyme activity was not due to an inhibitor in the tissue, nor is it due to a shift in the pH optimum. The deficiency of galactocerebroside beta-galactosidase as the primary enzymatic defect can account for the morphological and biochemical characteristics of this disease better than the previously reported deficiency of cerebroside-sulfatide sulfotransferase.

Adult↗
PubMed1971

On the control of the induction of -galactosidase in synchronous cultures of Escherichia coli.

The mode of induction of beta-galactosidase (EC 3.2.1.23) in synchronously growing cultures of two Hfr strains of Escherichia coli K12 was investigated. Cells can be induced to form the enzyme during any portion of the cell cycle; but when they are grown permanently in the presence of an inducer, enzyme synthesis is discontinuous. The interruption of beta-galactosidase synthesis appears to be geared to the growth cycle: it occurs when the cells are dividing actively. The observation that lac-specific messenger RNA is produced also in the absence of detectable enzyme synthesis suggests the existence of a control mechanism operating on the level of translation.

Bacterial Proteins↗
PubMed1974

Globoid cell leukodystrophy: deficiency of lactosyl ceramide beta-galactosidase.

Activity of lactosyl ceramide beta-galactosidase (beta-D-galactoside galactohydrolase, EC 3.2.1.23) was found to be extremely low in enzyme preparations from liver, brain, and cultured skin fibroblasts from patients with Krabbe's disease. Leukocytes from one set of parents had enzyme levels approximately half those measured in control leukocytes. The low activity observed for this galactolipid hydrolase is the fourth enzymatic deficiency noted for this genetic disease. Beta-galactosidase activity toward galactocerebroside, psychosine, and monogalactosyl diglyceride is also low in patients with Krabbe's disease. Other lysosomal enzymes measured were found to be in the normal range. This enzymatic defect may provide a better explanation for the pathological and chemical findings previously reported for this syndrome.

Adult↗
PubMed1997

Directed evolution of a fucosidase from a galactosidase by DNA shuffling and screening.

An efficient beta-fucosidase was evolved by DNA shuffling from the Escherichia coli lacZ beta-galactosidase. Seven rounds of DNA shuffling and colony screening on chromogenic fucose substrates were performed, using 10,000 colonies per round. Compared with native beta-galactosidase, the evolved enzyme purified from cells from the final round showed a 1,000-fold increased substrate specificity for o-nitrophenyl fucopyranoside versus o-nitrophenyl galactopyranoside and a 300-fold increased substrate specificity for p-nitrophenyl fucopyranoside versus p-nitrophenyl galactopyranoside. The evolved cell line showed a 66-fold increase in p-nitrophenyl fucosidase specific activity. The evolved fucosidase has a 10- to 20-fold increased kcat/Km for the fucose substrates compared with the native enzyme. The DNA sequence of the evolved fucosidase gene showed 13 base changes, resulting in six amino acid changes from the native enzyme. This effort shows that the library size that is required to obtain significant enhancements in specificity and activity by reiterative DNA shuffling and screening, even for an enzyme of 109 kDa, is within range of existing high-throughput technology. Reiterative generation of libraries and stepwise accumulation of improvements based on addition of beneficial mutations appears to be a promising alternative to rational design.

Escherichia coli↗
PubMed2000

Processing of lysosomal beta-galactosidase. The C-terminal precursor fragment is an essential domain of the mature enzyme.

Lysosomal beta-D-galactosidase (beta-gal), the enzyme deficient in the autosomal recessive disorders G(M1) gangliosidosis and Morquio B, is synthesized as an 85-kDa precursor that is C-terminally processed into a 64-66-kDa mature form. The released approximately 20-kDa proteolytic fragment was thought to be degraded. We now present evidence that it remains associated to the 64-kDa chain after partial proteolysis of the precursor. This polypeptide was found to copurify with beta-gal and protective protein/cathepsin A from mouse liver and Madin-Darby bovine kidney cells and was immunoprecipitated from human fibroblasts but not from fibroblasts of a G(M1) gangliosidosis and a galactosialidosis patient. Uptake of wild-type protective protein/cathepsin A by galactosialidosis fibroblasts resulted in a significant increase of mature and active beta-gal and its C-terminal fragment. Expression in COS-1 cells of mutant cDNAs encoding either the N-terminal or the C-terminal domain of beta-gal resulted in the synthesis of correctly sized polypeptides without catalytic activity. Only when co-expressed, the two subunits associate and become catalytically active. Our results suggest that the C terminus of beta-gal is an essential domain of the catalytically active enzyme and provide evidence that lysosomal beta-galactosidase is a two-subunit molecule. These data may give new significance to mutations in G(M1) gangliosidosis patients found in the C-terminal part of the molecule.

Animals↗
PubMed2001

Engineering regulable Escherichia coli beta-galactosidases as biosensors for anti-HIV antibody detection in human sera.

The activity of engineered, peptide-displaying enzymes is modulated by binding to specific anti-peptide antibodies. This new concept of a quantitative antibody detection system allows test kits to be set up for fast diagnosis of infectious diseases. To develop a quick and homogeneous assay for the detection of human immunodeficiency virus (HIV) infection, we have explored two acceptor sites of the bacterial Escherichia coli beta-galactosidase for the accommodation of HIV antigenic peptides. Two overlapping epitopes (namely P1 and P2) from the gp41 envelope glycoprotein, contained in different sized peptides, were inserted in the vicinity of the enzyme active site to generate a set of hybrid, enzymatically active beta-galactosidases. Regulable enzymes of different responsiveness to monoclonal antibody binding were generated with both acceptor sites tested. These biosensors were also sensitive to immune sera from HIV-infected patients. Modeling data provide insight into the structural modifications in the vicinity of the active site induced by peptide insertion that strongly affect the responsiveness of the engineered proteins through different parameters of their catalytic properties.

Amino Acid Sequence↗
PubMed2002

The Aes protein and the monomeric alpha-galactosidase from Escherichia coli form a non-covalent complex. Implications for the regulation of carbohydrate metabolism.

Aes, a 36-kDa acetylesterase from Escherichia coli, belongs to the hormone-sensitive lipase family, and it is involved in the regulation of MalT, the transcriptional activator of the maltose regulon. The activity of MalT is depressed through a direct protein-protein interaction with Aes. Although the effect is clear-cut, the meaning of this interaction and the conditions that trigger it still remain elusive. To perform a comparative thermodynamic study between the mesophilic Aes protein and two homologous thermostable enzymes, Aes was overexpressed in E. coli and purified. At the last step of the purification procedure the enzyme was eluted from a Mono Q HR 5/5 column as a major form migrating, anomalously, at 56 kDa on a calibrated Superdex 75 column. A minor peak that contains the Aes protein and a polypeptide of 50 kDa was also detected. By a combined analysis of size-exclusion chromatography and surface-enhanced laser desorption ionization-time of flight mass spectrometry, it was possible to demonstrate the presence in this peak of a stable 87-kDa complex, containing the Aes protein itself and the 50-kDa polypeptide in a 1:1 ratio. The homodimeric molecular species of Aes and of the 50-kDa polypeptide were also detected. The esterase activity associated with the 87-kDa complex, when assayed with p-nitrophenyl butanoate as substrate, proved 6-fold higher than the activity of the major Aes form of 56 kDa. Amino-terminal sequencing highlighted that the 50-kDa partner of Aes in the complex was the alpha-galactosidase from E. coli. The E. coli cells harboring plasmid pT7-SCII-aes and, therefore, expressing Aes were hampered in their growth on a minimal medium containing raffinose as a sole carbon source. Because alpha-galactosidase is involved in the metabolism of raffinose, the above findings suggest a potential role of Aes in the regulation of carbohydrate metabolism in E. coli.

Acetylesterase↗
PubMed2001

Improved method for the fluorimetric detection of beta-D-galactosidase in water.

A very convenient method to quantify coliform bacteria in water can probably be designed via the determination of the activity of the enzyme beta-D-galactosidase, whose natural occurrence is, apart from less frequently occurring aeromonads mainly restricted to this type of microorganisms. 4-methylumbelliferyl-beta-D-galactoside is used as substrate, which is hydrolyzed during the enzymatic reaction; the released 4-methylumbelliferone can be quantified fluorimetrically. In the present study the influence of various physical and chemical parameters on the determination is investigated and the experimental conditions are optimized. Most important entities are the pH value during hydrolysis, the presence of nutrients and co-factors in the sample, and the modification of the substrate. Statistical evaluation of the results obtained by changing single or multiple parameters reflects clearly their positive or negative influence on the enzyme activity. Thus, deliberate addition of surfactants, specific nutrients, salts and co-enzymes results in a significantly increased activity of beta-D-galactosidase towards the substrate, which can be advantageously exploited to increase the sensitivity of the analytical method together with a decrease of the detection limit. The influence of the parameters and the optimized conditions of the improved analytical methods are presented.

Enterobacteriaceae↗
PubMed2002

Effect of dietary supplementation with alpha-galactosidase preparation and stachyose on growth performance, nutrient digestibility and intestinal bacterial populations of piglets.

Two experiments were conducted to investigate the effect of dietary supplementation with alpha-galactosidase preparation and stachyose on growth performance, nutrient digestibility and intestinal bacterial populations of piglets. In Experiment I 72 crossbred piglets were allotted to three treatments, i.e. 1) control (basal) diet, 2) alpha-Gal1 and 3) alpha-Gal5, the basal diet supplemented with 0.01% and 0.05% alpha-galactosidase preparation (alpha-Gal), respectively. Average daily gain, average daily feed intake and feed conversion ratio were not influenced by alpha-Gal supplementation. In Experiment II 72 crossbred weanling piglets were randomly assigned to four treatments with a complete factorial design, i.e. 1) basal diet, 2) basal diet with 0.01% alpha-Gal supplementation, 3) basal diet with 1% stachyose supplementation and 4) basal diet with 1% stachyose and 0.01% alpha-Gal supplementation. Average daily gain and feed conversion ratio were influenced by alpha-Gal and stachyose supplementation. The ileal digestibility of stachyose, raffinose, gross energy and crude protein was improved significantly by alpha-Gal supplementation. The microbial populations in the intestine were modified by both alpha-Gal and stachyose supplementation. The data suggested that stachyose supplementation had an adverse effect on the growth performance of piglets and alpha-Gal supplementation could ameliorate it, especially in younger pigs.

Age Factors↗
PubMed1997

Purification, characterization, and expression of rat epididymal beta-D-galactosidase.

The purpose of the present study is to purify, kinetically characterize and measure the amount of soluble acid beta-D-galactosidase (EC 3.2.1.23) in different anatomical regions (caput, corpus, and cauda) of the adult rat epididymis. Based upon SDS-PAGE analysis, the subunit molecular mass of the caput and cauda enzyme is approximately 85,000 daltons while the corpus enzyme is approximately 50,000 daltons. The apparent Km and Vmax values are 67, 24, and 59 microM and 5.0, 1.88 and 6.3 microM/min./-mg protein for the enzyme purified from the caput, corpus, and cauda regions of the epididymis, respectively. However, no regional differences in the amount of soluble enzyme protein are observed. These data demonstrates regional differences in the activity of epididymal acid beta-D-galactosidase and suggest that the observed regional differences in enzyme activity may be due to posttranslational modifications.

Animals↗
PubMed2003

Comparison of chitin and Amberlite IRA-938 for alpha-galactosidase immobilization.

Watermelon alpha-galactosidase (EC 3.2.1.22) was immobilized on a natural (chitin) and a synthetic anion-exchange (Amberlite IRA-938) support by covalent coupling methods. The procedure entails the activation of supports with 1,1'-carbonyldiimidazole (CDI), followed by immobilization of the enzyme on to these supports without and with a spacer arm; gamma-aminobutyric acid (GABA). Optimization of activation was performed by changing the CDI concentrations and coupling efficiencies. The comparison of two immobilization techniques for both chitin and Amberlite IRA-938 was made by comparing different enzyme concentrations against enzyme activity yield. Furthermore, the storage stability of the immobilized enzymes was also investigated and chitin immobilized alpha-galactosidase was found to be better. Although the activity yield of immobilized enzymes were the same for both supports, the short storage stability of immobilized enzyme on Amberlite IRA-938 is currently a drawback to its applications.

Anion Exchange Resins↗
PubMed2004

Blood group B degrading activity of Ruminococcus gnavus alpha-galactosidase.

Ruminococcus gnavus is a Gram positive, nonspore-forming obligate anaerobe normally found in the human alimentary tract. In culture, this organism constitutively produces a 1-3 alpha-galactosidase. We fractionated and characterized this enzyme demonstrating hydrolysis of the B epitope on erythrocyte membranes and seroconversion to H epitope (blood type O). Since the enzyme yield was low, cell suspension studies could not be performed. Instead, hydrolysis of the B membrane epitope was studied with an ELISA. A highly purified enzyme product was analyzed for characteristics such as pH, ionic strength, and temperature optimum. Activity in red cell preservative solutions and in the presence of type B plasma was also demonstrated. Ruminococcus gnavus a 1-3 alpha-galactosidase has potential application in the enzymatic conversion of type B to O packed red blood cell units.

ABO Blood-Group System↗