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)

The proximal 5'-flanking region of the alpha-subunit gene from humans and cattle confers pituitary-specific expression to heterologous reporter genes in transgenic mice. To investigate whether these promoter regions also contain the necessary regulatory elements for cell-specific expression and hormonal regulation, we used three independent lines of transgenic mice. Two lines of transgenic mice contained chimeric genes consisting of either 1.6 kilobasepairs (kbp) of human or 3 15 basepairs of bovine alpha-subunit proximal 5'-flanking sequence linked to the bacterial gene encoding chloramphenicol acetyltransferase (CAT). A third line of transgenic mice contained the proximal 1.6 kbp of 5'-flanking sequence of the human alpha-subunit gene linked to the bacterial lacZ gene encoding beta-galactosidase (beta gal; H alpha beta gal transgenic mice). Hormonal replacement paradigms indicate that both human and bovine alpha CAT transgenes are regulated by GnRH, suggesting that their expression occurs in gonadotropes. Thus, the proximal 5'-flanking regions of both the human and bovine alpha-subunit genes must contain regulatory elements that confer both gonadotrope-specific expression and responsiveness to GnRH. In contrast to the human alpha-subunit promoter, the bovine alpha-subunit promoter lacks a functional cAMP response element, suggesting that transduction of both cell-specific and GnRH transcriptional signals occurs through cAMP response element-independent pathways. Thyrotropes also express the glycoprotein hormone alpha-subunit gene. Yet, hormone replacement paradigms with propylthiouracil and T3 were ineffective in altering CAT activity in the pituitary of human or bovine alpha CAT transgenic mice. Because a thyroid hormone response element has been localized to the proximal 5'-flanking region of the human alpha-subunit gene, these data suggest that the alpha CAT transgenes lack sufficient information to direct expression to thyrotropes. Direct evidence for this possibility was obtained through immunocytochemical studies performed on pituitaries from H alpha beta gal transgenic mice. beta-Galactosidase activity appeared in gonadotropes, but not thyrotropes. We conclude, therefore, that distinct and separable regulatory elements mediate the expression of the alpha-subunit gene in gonadotropes and thyrotropes.
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PMID:Gonadotrope- and thyrotrope-specific expression of the human and bovine glycoprotein hormone alpha-subunit genes is regulated by distinct cis-acting elements. 128 Mar 29

Myelin has pronounced effects upon the morphology, function, and growth of axons in the mammalian CNS. Consequently, oligodendrocyte development and myelination have been investigated using a wide variety of histological, immunocytochemical, ultrastructural, and biochemical techniques. While many of the spatial and temporal features of myelin appearance have been characterized, for any one species only limited regions of the CNS have been investigated. To address this limitation, we have derived transgenic mice in which the bacterial Lac Z gene is regulated by promoter elements of the myelin basic protein gene. When differentiating oligodendrocytes begin to elaborate recognizable myelin, they initiate expression of the MBP-Lac Z transgene and accumulate readily detectable levels of beta-galactosidase. Here, we exploit the sensitivity, resolution, and ease of beta-galactosidase histochemical assays to characterize the temporal and spatial patterns of CNS myelination in the mouse. Many features of the myelination program revealed by this approach were predicted by the immunocytochemical and ultrastructural data derived from other species. Nonetheless, previously undocumented patterns were also encountered. beta-Galactosidase was expressed first by oligodendrocytes in the ventral spinal cord, 1 d prior to birth. There, myelination proceeded in a strictly rostral-caudal direction, whereas in the dorsal cord, myelination initiated in the cervical enlargement and proceeded in both rostral and caudal directions. In the cerebellum, deep regions myelinated first, and in the optic nerve, myelination initiated at the retinal end. In contrast, the lateral olfactory tracts, pons, and optic chiasm initiated myelination along their entire course.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Myelin acquisition in the central nervous system of the mouse revealed by an MBP-Lac Z transgene. 128 97

beta-Galactosidase activity as illuminated by the indigogenic X-gal staining method has been used to demonstrate the presence of genetically modified cells carrying the reporter gene lacZ, coding for the E. coli enzyme. Endogenous activity has been assumed to be minimal since the pH optimum for the mammalian enzyme is 3.5-5.5, while the pH optimum for the E. coli enzyme (and thus of the staining procedure usually employed) is 7.3. Background staining has been reported to be limited to pericytes and a few specific neuronal cell groups. In contrast, our investigations of normal rat brain anatomy demonstrate that many specific neuronal cell groups possess endogenous beta-galactosidase activity when staining is performed at physiological pH. This suggests that background staining of endogenous beta-galactosidase activity in the rat brain has been underestimated. In addition, such specific activity would afford an additional means of identification and illustration of these cells.
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PMID:Demonstration of specific neuronal cell groups in rat brain by beta-galactosidase enzyme histochemistry. 128 64

Expression of the Vibrio fischeri luminescence genes (luxR and luxICDABEG) in Escherichia coli requires autoinducer (N-3-oxohexanoyl homoserine lactone) and LuxR protein, which activate transcription of luxICDABEG (genes for autoinducer synthase and the luminescence enzymes), and cyclic AMP (cAMP) and cAMP receptor protein (CRP), which activate transcription of the divergently expressed luxR gene. In E. coli and in V. fischeri, the autoinducer-LuxR protein-dependent induction of luxICDABEG transcription (called autoinduction) is delayed by glucose, whereas it is promoted by iron restriction, but the mechanisms for these effects are not clear. To examine in V. fischeri control of lux gene expression by autoinducer, cAMP, glucose, and iron, lux::Mu dI(lacZ) and lux deletion mutants of V. fischeri were constructed by conjugation and gene replacement procedures. beta-Galactosidase synthesis in a luxC::lacZ mutant exhibited autoinduction. In a luxR::lacZ mutant, complementation by the luxR gene was necessary for luminescence, and addition of cAMP increased beta-galactosidase activity four- to sixfold. Furthermore, a luxI::lacZ mutant produced no detectable autoinducer but responded to its addition with induced synthesis of beta-galactosidase. These results confirm in V. fischeri key features of lux gene regulation derived from studies with E. coli. However, beta-galactosidase specific activity in the luxI::lacZ mutant, without added autoinducer, exhibited an eight- to tenfold decrease and rise back during growth, as did beta-galactosidase and luciferase specific activities in the luxR::lacZ mutant and luciferase specific activity in a delta(luxR luxICD) mutant. The presence of glucose delayed the rise back in beta-galactosidase and luciferase specific activities in these strains, whereas iron restriction promoted it. Thus, in addition to transcriptional control by autoinducer and LuxR protein, the V. fischeri lux system exhibits a cell density-dependent modulation of expression that does not require autoinducer, LuxR protein, or known lux regulatory sites. The response of autoinducer-LuxR protein-independent modulation to glucose and iron may account for how these environmental factors control lux gene expressions.
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PMID:Cell density-dependent modulation of the Vibrio fischeri luminescence system in the absence of autoinducer and LuxR protein. 131 12

Examination of the role of carbohydrates in specific recognition between spermatozoa and zona pellucida has focussed on understanding the interaction of sperm hydrolases or lectin-like molecules with zona pellucida ligands. To elucidate the role of specific spermatozoan hydrolases in gamete interaction, rabbit testis beta-galactosidase and arylsulfatase A were purified, characterized, and localized in spermatozoa. beta-Galactosidase and arylsulfatase A co-purified after affinity, size, or reverse-phase chromatography. N-Terminal amino acid analysis and enzymatic characterization suggested that neither enzyme is a testis-specific isozyme. Size chromatography indicated that both enzymes aggregated into macromolecular complexes at pH 4.0, while both dissociated at pH 8.0. beta-Galactosidase and arylsulfatase A co-localized on the sperm surface and in the acrosome and postacrosomal regions of spermatozoa. Throughout the zona-induced acrosome reaction, both enzymes remained associated with the detached acrosomal cap and postacrosomal region of acrosome-reacted spermatozoa. Because the acrosome is an acidic subcellular compartment, internal beta-galactosidase and arylsulfatase A are probably aggregated in acrosome-intact spermatozoa and dissociate as they are exposed to pH increases during the acrosome reaction.
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PMID:Characterization of rabbit testis beta-galactosidase and arylsulfatase A: purification and localization in spermatozoa during the acrosome reaction. 135 47

We have constructed a shuttle plasmid for Bacillus megaterium and Escherichia coli that contains the promoter and repressor gene of the B. megaterium-borne operon for xylose utilization. A polylinker downstream of the promoter allows versatile cloning of genes under its transcriptional control. We have placed gdhA (encoding glucose dehydrogenase) from B. megaterium, lacZ (encoding beta-galactosidase) from E. coli, mro (encoding mutarotase) from Acinetobacter calcoaceticus, and human puk (encoding single-chain urokinase-like plasminogen activator, rscuPA) under xylose control in this vector. All four genes were between 130-fold and 350-fold inducible by 0.5% xylose in the growth medium in B. megaterium. Enzymatically active glucose dehydrogenase and mutarotase accumulated to 20% and 30% of the total soluble protein, respectively. beta-Galactosidase and rscuPA were also expressed at a high level. A gel analysis of the products demonstrated their proteolytic stability in the cytoplasm, even up to 5 h after induction. The expression properties of this new host-vector system are discussed in comparison to the ones available for B. subtilis and E. coli.
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PMID:Inducible high-level expression of heterologous genes in Bacillus megaterium using the regulatory elements of the xylose-utilization operon. 136 76

An experimental study was undertaken to evaluate alternative insect cell lines to Sf9 [from Spodoptera frugiperda (fall armyworm)] for the production of recombinant proteins. Insect cell lines from two different organisms were considered: IPLB-LdEIta (LdEIta) from Lymantria dispar (gypsy moth) and IPLB-HvT1 (HvT1) from Heliothis virescens (tobacco budworm). Both LdEIta and HvT1 produced higher total activity levels of recombinant beta-galactosidase in monolayer culture than Sf9 after infection with the Autographa californica nuclear polyhedrosis virus (AcMNPV). However, only LdEIta generated a product yield (activity per milligram of total protein) which exceeded that of Sf9 (by 25%), so its growth and production characteristics were investigated in depth. LdEIta generated production levels and yields of a recombinant rotaviral protein, VP4, which exceeded those of Sf9 by 84 and 38%, respectively. In suspension culture, the LdEIta cells grew as aggregates with a doubling time several hours longer than Sf9, but the recombinant product yields of LdEIta were still higher than Sf9 by 38% in this culture environment. beta-Galactosidase expression rates and cell death rates suggested that the difference in productivity between the two hosts was due to the ability of LdEIta to survive the baculovirus infection and produce recombinant proteins longer than Sf9. The presence of LdEIta aggregates in suspension culture may be used as a method to separate live cells from dead cells, labile product, and spent medium in recombinant protein production processes.
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PMID:Production of recombinant proteins by baculovirus-infected gypsy moth cells. 136 95

Several nitrofurans were found to induce umu gene expression in Salmonella typhimurium TA1535/pSK1002 as defined on the basis of at least a 2-fold increase of beta-galactosidase activity over the background level. beta-Galactosidase activity increased with increasing concentrations of the chemical, attained a maximum at a concentration which was different for different nitrofurans used, and then gradually decreased with a further increase of the nitrofuran concentration. The umu gene expression test revealed that the genotoxic activity was highest for furazolidone and lowest for 5-nitro-2-furaldehyde.
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PMID:On the induction of umu gene expression in Salmonella typhimurium strain TA1535/pSK1002 by some nitrofurans. 137 46

A plasmid shuttle vector (pSP10) was designed and constructed to simplify screening of cloned DNA and to facilitate expression of the protein products. The plasmid contained the following features: (i) a selection gene, chloramphenicol acetyltransferase; (ii) an indicator gene encoding beta-galactosidase for visual identification of colonies containing DNA inserts; (iii) a cloning region immediately upstream from the indicator gene; (iv) origins of replication recognized by both Escherichia coli and Bacillus subtilis; and (v) a synthetic DNA expression control sequence, including -35 and -10 regions, ribosomal binding site, and transcriptional and translational start sites. The promoter region is a synthetic consensus sequence derived from published B. subtilis promoters. The plasmid has been shown to replicate actively in E. coli and B. subtilis and to confer chloramphenicol resistance to both hosts. DNA inserted at the cloning region inactivates the indicator gene, resulting in white colonies on 5'-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside plates. beta-Galactosidase has been expressed from pSP10 in both E. coli and B. subtilis. A comparison was made of the expression levels of beta-galactosidase from the same plasmid which had been modified to contain: (i) the synthetic control region, (ii) no promoter region, (iii) the synthetic control region cloned in the opposite orientation, or (iv) the tac promoter.
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PMID:Protein expression from an Escherichia coli/Bacillus subtilis multifunctional shuttle plasmid with synthetic promoter sequences. 139 13

ConA was immobilized on an epoxy-activated copolymer of 2-hydroxyethyl-methacrylate and ethylene-dimethacrylate and commercially available high-pressure liquid chromatography (HPLC) sorbents Separon HEMA 1000 EL, Separon HEMA 1000 E, and Separon HEMA 1000 EH (Tessek, Prague, CSFR Denmark). Specific, sensitive, and rapid method for determination of immobilized ConA lectin activity was developed. beta-Galactosidase from Aspergilus oryzae oligomannosyl residues was used as specific affinant. After separation of bound and unbound beta-galactosidase, enzyme activity was measured in supernatant and thus immobilized ConA lectin activity was calculated easily. The use of the method for evaluating the properties of immobilized ConA, efficiency of immobilization, specific activity, and thermostability is shown. The method developed could be generalized by using artificially glycosylated enzyme for any lectin.
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PMID:Rapid determination of immobilized ConA lectin activity. 141 45


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