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 merodiploids of Escherichia coli that contain genes for the lac operon on both chromosome and episome were tested for production of lac enzymes after growth on various carbon sources. The specific activity of beta-galactosidase (and of thiogalactoside transacetylase) was about twice that from haploid cells when grown on glycerol. With succinate as carbon source, the specific activity increased by an additional factor of 3. Up to 25% of the soluble cell protein is beta-galactosidase in these strains, one of which is inducible and the other constitutive. The enzyme is purified easily in high yield by ammonium sulfate fractionation and electrophoresis.
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PMID:High-level production of -galactosidase by Escherichia coli merodiploids. 456 80

A method is described for location of proteins in bacteria. It depends upon two techniques. One technique is the inactivation of the protein by a reagent which is incapable of penetrating the bacterial membrane (permeability barrier). Proteins inside this membrane cannot be inactivated unless the cells are disrupted; proteins on or outside the membrane can be inactivated. The second technique depends upon inactivation of the protein by specific antibody. Antibody should not penetrate the external bacterial wall, and therefore should only inactivate proteins that are on the wall surface. Thus, proteins can be localized inside the membrane, in the wall-membrane area, or outside the wall. One reagent developed for use with the first technique is diazo-7-amino-1,3-naphthalene-disulfonate. It inactivated beta-galactoside transport, but not beta-galactosidase of intact Escherichia coli. Similarly, it inactivated sulfate binding and transport but not uridine phosphorylase activity of Salmonella typhimurium. This indicates that the sulfate-binding protein is on or outside the cell membrane, and that uridine phosphorylase is inside the cell. The organic mercurial compounds used also showed that the sensitive parts of the sulfate and alpha-methylglucoside transport systems are less reactive than the sensitive part of the beta-galactoside system. Antibody to the sulfate-binding protein inactivated the purified protein but did not inactivate this protein when intact bacteria were employed. Thus, it appears that the sulfate-binding protein does not protrude outside the cell wall. The conclusion that the binding protein is located in the wall-membrane region is supported by its release upon spheroplast formation or osmotic shock, and also by its ability to combine with sulfate in bacteria which cannot transport sulfate into the cell.
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PMID:Location of sulfate-binding protein in Salmonella typhimurium. 497 82

beta-Galactosidase of Streptococcus lactis 7962 was partially purified, and its properties were studied. Enzyme from only this strain of numerous lactic streptococci tested was stable in cell exudates prepared by various means. Cell-free extracts of the 7962 strain were prepared by sonic treatment of washed cells previously grown in presence of lactose to fully induce enzyme synthesis. Protamine sulfate precipitation of the nucleic acids and ammonium sulfate precipitation of protein were used for partial purification of the enzyme. The resulting enzyme, when resuspended in cold (5 C) phosphate buffer, was extremely labile. However, ammonium sulfate in high concentrations (0.85 m) stabilized and stimulated beta-galactosidase activity. Sephadex G-200 gel filtration was used to achieve further purification and to monitor homogeneity of the enzyme. Separation of the beta-galactosidase in buffer at 5 C yielded an enzyme elution pattern showing two peaks of activity. However, addition of the enzyme solution in 0.85 m ammonium sulfate to the column equilibrated with the same salt concentration yielded only one peak of enzyme activity. The data suggested that the native enzyme was dissociating into active subunits which were stabilized in the presence of the ammonium sulfate.
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PMID:Purification and properties of Streptococcus lactis beta-galactosidase. 602 31

We describe the use of a combined cloning/expression protocol to identify a gene encoding a Pseudorabies virus (PRV) glycoprotein. Prior to this study, the genome locations of PRV glycoproteins had not been described. We first identified PRV glycoproteins using antibodies directed against PRV virions. Using affinity chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the PRV glycoproteins were separated and antibodies were made against them in rabbits. Using DNase digestion, PRV genomic DNA was fragmented into approximately 500-base-pair regions. These random fragments were inserted in an expression vector and the PRV DNA sequences were expressed as proteins fused to beta-galactosidase. The antibodies made in rabbits were then used as probes in a Western blot analysis to screen for the presence of PRV-specific glycoprotein sequences in these PRV-beta-galactosidase fusion proteins. Two expression plasmids were isolated that specified fusion proteins that reacted in the Western blot analysis with rabbit antibodies directed against a 74,000 molecular weight PRV glycoprotein. The PRV DNA sequences represented in the expression plasmids were mapped within a single BamHI fragment of PRV genomic DNA, but were not overlapping. PRV-beta-galactosidase fusion proteins produced by these two expression plasmids were used to inoculate rabbits. Antibodies produced against both fusion proteins recognized PRV-specific glycoproteins of 74,000 and 92,000 molecular weight. This protocol should have general application in localizing genes within large DNA virus genomes.
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PMID:Construction of E. coli expression plasmid libraries: localization of a pseudorabies virus glycoprotein gene. 609 May 66

Cerebroside sulfatase (CSase) activator was isolated from human liver by acetone precipitation, anion-exchange chromatography, gel filtration and polyacrylamide gel electrophoresis. The CSase activator was a heat-stable protein with an isoelectric point of 4.54. Molecular weight (Mr) of the activator was estimated as 22,000 with the gel permeation and about 8,000 by gel electrophoresis in the presence of sodium dodecyl sulfate, suggesting that the native activator is a trimer of a subunit with Mr 8,000. The CSase activator formed a complex with an equimolar amount of cerebroside sulfate (CS), when examined by gel permeation experiments. The activator also bound to galactosylceramide and GM2 ganglioside but scarcely to GM1 ganglioside, and activated to some extent beta-N-acetyl-hexosaminidase A and beta-galactosidase, although the CSase activator could be clearly distinguished from the GM1 beta-galactosidase activator so far known. Though the affinity chromatography using glycolipid ligands, the CSase activator did not recognize sulfate group of CS, but appeared to have a relatively broad specificity for lipid-linked hexose.
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PMID:[Purification and characterization of cerebroside sulfatase activator]. 614 Nov 30

The mechanism by which large premature termination fragments of beta-galactosidase were degraded in Escherichia coli was studied using quantitative immunoprecipitation techniques. Two different lacZ nonsense mutants which produced apparent primary translation products of 96,000 and 109,000 daltons, respectively, were both shown to produce a second beta-galactosidase-related polypeptide of Mr = 90,000. These 90,000-dalton polypeptides appeared to be the same in both strains since they co-migrated when analyzed as a mixture on sodium dodecyl sulfate-polyacrylamide gels and were indistinguishable when analyzed by one-dimensional peptide mapping. Pulse-chase experiments established a stoichiometric precursor-product relationship between the primary mutant gene products (called the A polypeptides) and the common 90,000-dalton polypeptide (called the B polypeptide). No intermediates were detected between the A and B polypeptides. We propose that there is a common pathway for the degradation of these different large fragments of beta-galactosidase. According to this model, the first step would be a specific endoproteolytic cleavage of the primary translation product which produces the 90,000-dalton polypeptide as a common intermediate. The kinetic analysis demonstrated a first order decay of both A and B polypeptides but, surprisingly, the first order rate constant for the decay of A appeared dependent upon the induction regimen. This result suggested that degradation may possibly be autoregulated either by the intracellular level of A or by other intermediates in the degradation pathway.
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PMID:Limited proteolysis. Early steps in the processing of large premature termination fragments of beta-galactosidase in Escherichia coli. 616 13

Inhibition by low-molecular-weight sugars of precipitin line formation between a polysaccharide (EF) excreted by Leishmania tropica subsp. major, Leishmania enriettii, and rabbit antileishmanial antibodies on double gel diffusion plates revealed that galactose residues, possibly as components of lactosyl groups, were the critical immunodominant sugars mediating antibody recognition of EF. The galactose residues of the EF of L. tropica subsp. major were specifically labeled with tritium via galactose oxidase and sodium boro[3H]hydride. The radioactive EF had an apparent molecular weight of about 85,000 on sodium dodecyl sulfate-polyacrylamide gels and was precipitated by antileishmanial antibodies as well as Ricinus communis lectins I and II (galactose specific). Lectins specific for glucose-mannose residues, fucose, N-acetylglucosamine, and N-acetylgalactosamine did not precipitate the labeled EF. Treatment of [3H]EF with proteolytic (trypsin, papain, protease) or glycosidic (alpha-amylase, beta-galactosidase) enzymes had no effect on either the electrophoretic pattern of the material or on its recognition by antileishmanial antibodies or R. communis lectin. This resistance to enzyme activity suggests that EF may be a useful marker for the presence of the parasite in vivo if it can be detected in minute quantities.
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PMID:Identification of galactose as the immunodominant sugar of leishmanial excreted factor and subsequent labeling with galactose oxidase and sodium boro[3H]hydride. 617 74

An 18-year-old boy showed childhood onset of mental retardation, neurogenic muscle atrophy with hyperreflexia, Marfan-like features, multiple epiphyseal dysplasia, increased urinary excretion of dermatan sulfate, and decreased lysosomal enzyme activities in beta-galactosidase, beta-glucuronidase, and N-acetyl-beta-D-glucosaminidase. This case may be a new syndrome, the combination of neurogenic muscle atrophy with lysosomal enzyme deficiencies.
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PMID:Juvenile neurogenic muscle atrophy with lysosomal enzyme deficiencies: new disease or variant of mucopolysaccharidosis? 618 76

A fetal antigen (FA) was isolated from spent culture medium of a melanoma (M14) cell line. Allogeneic serum samples from melanoma patients, previously characterized with respect to anti-FA activity, were used as the source of anti-FA antibody. The FA activity was partially purified by membrane ultrafiltration, gel filtration, and chloroform:methanol extraction. The partially purified FA was then used to develop an enzyme-linked immunosorbent assay (ELISA). By indirect ELISA both the IgG and IgM classes of anti-FA antibodies were detected in the sera of cancer patients and normal volunteers. The incidences of anti-FA antibodies in the sera of cancer patients and normal volunteers were not significantly different. As detected by competitive inhibition in ELISA, FA activity was widely distributed among melanoma, sarcoma, and carcinoma tumor tissues and cultured tumor cells, as well as among fetal brain, skin, and muscle tissues. FA activity was destroyed by treatment with beta-galactosidase and hyaluronidase, but it was not destroyed by proteolytic and lipolytic enzymes. The antigen bound to immobilized ricin, peanut, and soybean lectins. FA activity in material purified by ricin-affinity chromatography was associated with molecules in the 60,000- to 70,000-dalton region as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These results suggest a glycoprotein nature for the FA isolated from the spent culture medium of melanoma (M14) cells; this FA apparently elicits formation of natural antibodies in the cancer patients and normal donors.
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PMID:Immunochemical characterization of fetal antigen isolated from spent medium of a human melanoma cell line. 619 35

beta-Galactosidase purified to apparent homogeneity from human placenta occurred in two separable fractions. A low molecular mass form (relative mass (Mr) 170 000) is composed of a single polypeptide chain (Mr 70 000). This was derived from a larger form by molecular sieve chromatography at both low (10 mM) and high (500 mM) NaCl concentration. The larger form of beta-galactosidase also contains small amounts of two polypeptides with apparent Mr values of 23 000 and 35 000 daltons. Both forms of the enzyme hydrolyze synthetic aryl galactosides and natural glycolipid substrates at comparable rates. Antibodies raised in rabbits against the low Mr beta-galactosidase also cross-reacts with the high Mr enzyme. The antibody preparation also cross-reacted with beta-hexosaminidase even though the latter was found at very low levels in the antigen, as judged by lack of detection of representative protein bands on sodium dodecyl sulfate - polyacrylamide gel electrophoresis and enzyme activity measurements. A portion of this cross-reactivity (35%) against beta-hexosaminidase could not be absorbed from the preparation without the simultaneous loss of beta-galactosidase activity, suggesting that the two enzymes show a degree of antigenic identity.
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PMID:Human placental beta-galactosidase: structural and immunological observations. 620 36


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