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)

We have isolated a novel cDNA from Euglena gracilis that encodes a protein composed of 24.9% aspartate with an estimated pI of 3.56, and a deduced molecular mass of 73,542 Da. The first 20 or so amino acids are hydrophobic and resemble a signal sequence. The rest of the polypeptide is composed of a 23-amino-acid repeat. There are 30 repeats, of which 23 are full length. Part of the consensus sequence derived from the repeats has some similarity to the loop of the EF-hand type calcium-binding motif. Evidence is presented that a fusion protein of this novel protein with beta-galactosidase can bind calcium. Northern blotting indicates a single transcript of 2.3 kb (the same size as the cDNA). In-vitro translation of the cDNA gives a protein that migrates on SDS/PAGE with an apparent molecular mass of 120-125 kDa. The protein is processed into a smaller, protease-protected form (110-120 kDa) when translated in the presence of canine pancreatic microsomal vesicles. This suggests that the protein is targeted across the endoplasmic reticulum membrane in vivo, and is the first report of a signal sequence from E. gracilis. We propose that the cDNA obtained encodes a novel calcium-binding protein that is either secreted or resident in the endomembrane system of E. gracilis, and call it the acidic-repeat protein.
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PMID:A novel calcium-binding protein from Euglena gracilis. Characterisation of a cDNA encoding a 74-kDa acidic-repeat protein targeted across the endoplasmic reticulum. 128 88

We have characterized two mutants of Rhizobium meliloti L5-30 obtained by random mutagenesis using Mu-lacZ that were defective in transport of C4-dicarboxylic acids. These mutants induced ineffective nodules on alfalfa. Mutations in the two strains appeared to be located in a dctA gene. Levels of dctA gene expression were determined under different environmental conditions using dctA-lacZ fusions, and beta-galactosidase activities increased in response to osmotic stress and also when the cells were incubated in medium low in calcium. The transcriptional induction of the dctA gene by environmental signals was decreased by DNA gyrase inhibitors such as novobiocin and coumermycin.
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PMID:Induction of C4-dicarboxylate transport genes by external stimuli in Rhizobium meliloti. 131 20

Northern-blot analysis was used to demonstrate that an increase in extracellular glucose concentration increased the content of preproinsulin mRNA 2.3-fold in the beta-cell line HIT T15. A probe for the constitutively expressed glyceraldehyde-3-phosphate dehydrogenase was used as a control. Mannoheptulose blocked this effect of glucose. A stimulatory effect on preproinsulin mRNA levels was also observed in response to mannose and to 4-methyl-2-oxopentanoate. However, galactose and arginine were ineffective. Glucagon, forskolin and dibutyryl cyclic AMP also elicited an increase in HIT-cell preproinsulin mRNA. The ability of the 5' upstream region of the preproinsulin gene to mediate the effect of glucose and other metabolites on transcription was studied by using a bacterial reporter gene technique. HIT cells were transfected with a plasmid, pOK1, containing the upstream region of the rat insulin-1 gene (-345 to +1) linked to chloramphenicol acetyltransferase (CAT). Co-transfection with a plasmid pRSV beta-gal containing beta-galactosidase driven by the Rous sarcoma virus promoter was used as a control for the efficiency of transfection; expression of CAT activity in transfected HIT cells was normalized by reference to expression of beta-galactosidase. Glucose caused a dose-dependent increase in expression of CAT activity, with a half-maximal effect at 5.5 mM and a maximum response of 4-fold. Mannoheptulose blocked this effect of glucose. Other metabolites (mannose, 4-methyl-2-oxopentanoate and leucine plus glutamine) were also able to increase insulin promoter-driven CAT expression, but galactose and arginine were ineffective. The stimulatory effect of glucose on CAT expression was not blocked by verapamil and was inhibited by increasing extracellular Ca2+ from 0.4 to 5 mM. Both dibutyryl cyclic AMP and forskolin caused an increase in insulin promoter-driven gene expression in the presence of 1 mM-glucose, but neither agent further increased the level of expression occurring in the presence of a maximally stimulating glucose concentration. The phorbol ester phorbol 12-myristate 13-acetate (PMA) also increased insulin promoter-driven CAT expression in the presence of 1 mM-, but not 11 mM-glucose. Staurosporine blocked the stimulatory effect not only of PMA but also of glucose and of dibutyryl cyclic AMP. We conclude that the 5' upstream region of the insulin gene contains sequences responsible for mediating the stimulatory effect of glucose on insulin-gene transcription.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Control of insulin gene expression by glucose. 132 37

The stable transfection of immortalized Rat-1 and rat skin primary fibroblast cell lines by calcium phosphate precipitation, lipofection and electroporation methods have been examined. The lipofection method was found to be better than the other methods in terms of higher transfection efficiency and convenient use. Expression of beta-galactosidase from two different viral promoters showed that the level of transgene expression depends on the promoter strength in a particular cell type. The results presented here show that the transgene expression is extremely variable among different colonies generated from individually transfected cells. Therefore, it is necessary to examine individual colonies of cells for the production of reporter gene to obtain cell lines expressing high amounts of gene products.
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PMID:Gene transfer into established and primary fibroblast cell lines: comparison of transfection methods and promoters. 133 35

A reaction-diffusion model was used to simulate a co-immobilized system utilizing the numerical method of orthogonal collocation. The production of ethanol from deproteinized whey using beta-galactosidase co-immobilized with Saccharomyces cerevisiae in calcium alginate gel beads was chosen as a model system. Calculated concentrations of lactose, glucose, galactose and ethanol were compared with experimental data for a batch reactor and a continuous horizontal packed-bed reactor. The mathematical model has been used to analyse the influence of internal and external mass transfer for the continuous reactor. The external mass transfer was shown to be of minor importance. The introduction of baffles decreased the backmixing in the horizontal packed-bed reactor. Internal mass transfer was found to be the main cause of the reduction in the apparent reaction rate. Thus, much of the expected increase in reaction rate is diminished by mass transfer hindrance when the cell concentration is increased.
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PMID:Simulation of batch and continuous reactors with co-immobilized yeast and beta-galactosidase. 136 20

Tammar wallaby (Macropus eugenii) mammary glands contain a UDP-GlcNAc:Gal beta 1----3Gal beta 1----4Glc beta 1----6-N-acetylglucosaminyltransferase (GlcNAcT) whose activity has been characterized with respect to the effect of pH, apparent Km for acceptor, effects of bivalent metal ions, acceptor specificity and identity of products. The enzyme did not show an absolute requirement for any bivalent metal ion but its activity was increased markedly by Mg2+, Ca2+ and Ba2+ and, to a lesser extent, by Mn2+. When Gal beta 1----3Gal beta 1----4Glc was used as acceptor, the product was Gal beta 1----3[GlcNAc beta 1----6]Gal beta 1----4Glc. With Gal beta 1----3Gal beta 1----3Gal beta 1----4Glc as acceptor, the product was shown, by 1H-NMR spectroscopy and exo-beta-galactosidase digestion, to be a novel pentasaccharide with the structure Gal beta 1----3[GlcNAc beta 1----6]Gal beta 1----3Gal beta 1----4Glc, suggesting that the enzyme recognises the non-reducing end of the acceptor substrate, rather than the reducing end.
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PMID:Biosynthesis of marsupial milk oligosaccharides. II: Characterization of a beta 6-N-acetylglucosaminyltransferase in lactating mammary glands of the tammar wallaby, Macropus eugenii. 138 55

Transfection of the beta-galactosidase gene in quiescent cultures of adult rat hepatocytes with the calcium phosphate precipitate or the lipofection methods gave a higher level of beta-galactosidase gene expression with the lipofection than with the calcium phosphate precipitate method, but the transfection efficiency was weak in both cases. Transfection of hepatocytes stimulated to proliferate before transfection either in vivo by partial hepatectomy or in vitro by epidermal growth factor was more efficient than transfection of quiescent hepatocytes, and the lipofection method gave better results than the calcium phosphate precipitate method.
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PMID:Increased efficiency of gene transfection in primary cultures of adult rat hepatocytes stimulated to proliferate: a comparative study using the lipofection and the calcium phosphate precipitate methods. 151 43

The main objective of the study was to investigate the effect of the calcium antagonists Verapamil (2 mg.kg-1.day) and Nifedipine (1 mg.kg-1.day-1) on cholesterol (1%) induced atherosclerosis in rabbits. The drugs were administered s.c. twice daily over a period of 8 weeks. Blood lipid levels were determined three times during the experiment. After the experimental period the animals were killed and macroscopic changes on the aorta were recorded. For histochemical investigation samples were taken from the arch of the aorta and coronary artery. In cryostat sections lipids were determined by Sudan black B and Fett rot 7 B and the following enzymes were assayed: acid phosphatase, non-specific and acid esterase, acid beta-galactosidase, dipeptidyl peptidase I and II, and glucose-6-phosphate dehydrogenase. Following treatment with the calcium antagonists the levels of triacylglycerols and of total cholesterol were significantly increased in comparison with the control and diet groups. The ratio of HDL cholesterol to total cholesterol decreased in the treated animals. In lipoid plaques the activity of enzymes was enhanced in all experimental animals. There were however no qualitative differences in the composition of plaques between individual groups, which exhibited only quantitative differences. The number of migrating macrophages was increased only in the nifedipine treated animals. The extent of plaques was significantly decreased after nifedipine treatment, whereas verapamil failed to exert antiatherogenic effect. (Tab. 2, Fig. 4, Ref. 22.).
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PMID:[The effect of verapamil and nifedipine on the development of experimental atherosclerosis in rabbits]. 152 79

Mammalian cell lysate containing beta-galactosidase (beta Gal) derived from the transient expression of the bacterial lacZ gene driven by the human beta-actin promoter loses activity progressively over time in storage at -20 degrees C in the presence of EDTA. The simultaneous presence of NaCl with EDTA exacerbates such an inactivation, although NaCl by itself does not. However, EGTA, a chelating agent that preferentially binds Ca2+ over Mg2+, does not inactivate beta Gal. Addition of equal or higher molar concentration of Mg2+ (as MgCl2) or Ca2+ (as CaCl2), both effectively chelated by EDTA, to an EDTA-containing lysate prevents this cold-related inactivation, but does not reactivate the enzyme. Therefore, the chelation of Mg2+ by EDTA at -20 degrees C inactivates beta Gal. Storage of cell lysate at -70 degrees C completely prevents the EDTA-induced inactivation of beta Gal. It is recommended that when beta Gal activity is used as the reporter for gene expression 1) EDTA should not be used to prepare cell lysate and 2) the cell lysate should be stored in a -70 degrees C freezer to preserve full activity.
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PMID:Magnesium chelation inactivates beta-galactosidase in -20 degrees C storage. 161 4

A human hippocampus cDNA library in lambda ZAP II was screened by hybridization with a rat brain inositol 1,4,5-trisphosphate (InsP3) 3-kinase cDNA. Two clones (hh6 and hh3) were isolated and sequenced. The insert of clone hh6 was shown to correspond to the 3' end of the coding sequence of 50,000-Mr InsP3 3-kinase (referred to as 3-kinase-A). Sequencing of the clone hh3 insert yielded an open reading frame encoding a 472-amino acid protein with a calculated Mr of 53,451 (referred to as 3-kinase-B). The C-terminal part of 3-kinase-B (residues 187-462) was 68% identical with 3-kinase-A in amino acid sequence. The cDNA of clone hh3 was rescued as a Bluescript plasmid and expressed in Escherichia coli as a beta-galactosidase fusion product. It showed InsP3 3-kinase activity that was stimulated in the presence of Ca2+/calmodulin (more than 7-fold in a crude bacterial lysate from expressed plasmid). Regeneration of InsP3 3-kinase activity after SDS/PAGE identified a major polypeptide (Mr 60,000-65,000). The Km for InsP3 of expressed 3-kinase-B was 1.6 microM. These data provide molecular evidence for the existence of InsP3 3-kinase isoenzymes.
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PMID:Molecular cloning and expression of a new putative inositol 1,4,5-trisphosphate 3-kinase isoenzyme. 165 94


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