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)

The retrograde transneuronal viral tracing method was used to study the CNS nuclei that innervate the parasympathetic preganglionic neurons controlling the submandibular gland in the rat. A genetically engineered beta-galactosidase expressing Bartha strain of pseudorabies virus (PRV) was injected into the submandibular gland of rats. After 4 days, PRV infected tissues were reacted with the Bluo-Gal substrate (halogenated indolyl-beta-D-galactoside) and labeled cell bodies were identified throughout the brain. In the medulla oblongata, cell body labeling was seen in the superior salivatory nucleus, and throughout the medullary reticular formation as well as in the nucleus of the solitary tract, spinal trigeminal nucleus, and deep cerebellar nuclei. In the pons, PRV labeled neurons were found bilaterally in the locus ceruleus, subceruleus region, and parabrachial complex. In the mesencephalon, labeled cells were found in the Edinger-Westphal nucleus, deep mesencephalic nucleus, and central grey matter. Several hypothalamic regions were labeled including the lateral, perifornical and paraventricular hypothalamic nuclei. In the telencephalon, PRV-positive cell bodies were observed in the substantia innominata, bed nucleus of the stria terminalis and central nucleus of the amygdala. The results suggest that widespread areas of the CNS are involved in control of salivation.
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PMID:CNS cell groups projecting to the submandibular parasympathetic preganglionic neurons in the rat: a retrograde transneuronal viral cell body labeling study. 131 71

In order to define the cellular specificity of the interphotoreceptor retinoid-binding protein (IRBP) promoter in the retina, we linked the human IRBP promoter to the beta-galactosidase (lacZ) gene and made five lines of transgenic mice. In three of the five transgenic mouse lines, retinas showed positive staining upon incubation with 5-bromo-4-chloro-3-indolyl-beta-D-galactoside (X-gal). Mice from one line (OVE278B) showed positive X-gal staining throughout the retina except for the most peripheral regions. Interestingly, the staining was heterogeneous throughout the retina. Heavily stained regions were interspersed with lightly stained areas. Mice in two other lines showed highly mosaic X-gal staining patterns. Histological examination demonstrated that staining was confined to photoreceptor cells in all three expressing families. Furthermore, electron microscopy showed that the promoter is active in both rod and cone cells. Our results demonstrate that the human IRBP promoter can be used to obtain photoreceptor-specific gene expression in transgenic mice.
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PMID:Photoreceptor-specific activity of the human interphotoreceptor retinoid-binding protein (IRBP) promoter in transgenic mice. 142 58

Free biotin was quantitated by a competition by coating biotin-bovine serum albumin conjugate on a polystyrene microplate for binding to avidin-beta-galactosidase conjugate. The enzyme conjugate remaining on the plate surface as a result of the competition was detected by reaction with one of the following fluorogenic substrates, resorufin beta-D-galactoside and fluorescein di-beta-D-galactoside, in a fluorescence plate reader. Free biotin as little as 0.1 nmol can be routinely detected.
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PMID:A simple and sensitive enzyme-mediated assay of biotin. 147 23

Salmonella typhimurium has the capacity to enter into and multiply within epithelial cells. During the entire intracellular stage, bacteria are enclosed within a vacuole. To characterize the micro-environment of the bacteria-containing vacuoles, we have used a new method to measure the expression levels of several S. typhimurium genes in intracellular bacteria within Madin-Darby canine kidney (MDCK) epithelial cells. Our study was based on the determination of beta-galactosidase activity derived from lacZ transcriptional fusions using the highly sensitive substrate fluorescein-di-beta-D-galactoside (FDG). Expression of the iroA and mgtB genes (induced by Fe2+ and Mg2+ limitation respectively), and cadA (induced by pH 6.0 in the presence of lysine, with enhanced expression under anaerobiosis) were characterized at different post-infection times. High intracellular expression levels were detected for the iroA and mgtB genes, suggesting that the concentrations of free Fe2+ and Mg2+ in the vacuole may be low. cadA activity was detected only at early post-infection times (4 h), suggesting that the vacuole may have a mild-acidic pH, and oxygen and lysine present at this time. Globally, the results reported indicate that the use of a highly sensitive beta-galactosidase substrate can provide information about the micro-environment within which an intracellular pathogen, such as S. typhimurium, resides.
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PMID:Characterization of the micro-environment of Salmonella typhimurium-containing vacuoles within MDCK epithelial cells. 148 85

The mutational potency of apurinic/apyrimidinic (AP) sites induced by heat-treatment under acidic conditions has been studied in mammalian cells. Abasic sites were induced on a single-stranded DNA shuttle vector carrying the supF tRNA gene, eliminating, therefore, any ambiguity concerning the damaged strand. This vector was able to replicate both in mammalian cells and in bacteria where the mutations induced in animal cells on the supF tRNA gene were screened by the white/blue beta-galactosidase assay in the presence of isopropyl-1-thio-beta-D-galactopyranoside and 5-bromo-4-chloro-3-indoyl-beta-D-galactoside. All white colonies contained plasmid with a mutation on the target gene which was directly sequenced. Our results show that one AP site was induced/22 min of heating as measured by sensitivity of DNA to alkali denaturation or treatment with the AP-endonuclease activity of the FPG protein (Fapy-DNA glycosylase). Putative AP sites decrease survival of the plasmid with a lethal hit of one AP site/single-stranded molecule. Mutation frequency was increased by a factor of approximately six after 2 h at 70 degrees C. Most of the induced mutations were point mutations not distributed at random and clustered in the gene region which will give rise to the mature tRNA. Mutations were abolished by treatments that eliminated AP sites such as alkali treatment or incubation with the Fapy-DNA glycosylase protein. Under our experimental conditions, when only single mutations were taken into account, the order of base insertion opposite AP sites was G greater than A greater than T greater than C.
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PMID:Mutation spectrum of heat-induced abasic sites on a single-stranded shuttle vector replicated in mammalian cells. 152 92

In the present report, we describe the establishment of a cell line that can be used as the target for measuring the activity of cytotoxic T lymphocytes (CTL) by an enzyme release assay. We transfected P3/NS1-Ag4-1 (NS-1), a myeloma cell line derived from BALB/c mice with Escherichia coli beta-galactosidase (beta-Gal) gene, and isolated a stable transformant designated as NS-1/Z that expressed a high level of the enzyme activity intracellularly. The effector cells showing cytotoxicity against NS-1/Z were induced when the spleen cells of AKR or C3H mice were cultured with mitomycin C-treated BALB/c spleen cells for 4 days. When 2 x 10(4) NS-1/Z cells were incubated with varying numbers of effector cells, beta-Gal activity was released from the target cells depending on the number of effector cells and the time of incubation for up to 8 h. A highly sensitive enzyme assay was performed by using a fluorescent substrate, 4-methylumbelliferyl-beta-D-galactoside. The cytotoxicity was specific for H-2 haplotype of the stimulator cells, and was abolished by treating the effector cells with anti-Lyt 2 plus complement. The sensitivity of the enzyme release assay was comparable to that of 51Cr release assay. These results indicate that NS-1/Z can be used as a target cell line for the non-radioactive measurement of CTL activity.
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PMID:Establishment of an enzyme release assay for cytotoxic T lymphocyte activity. 154 35

A beta-galactosidase expression pseudorabies virus (Bartha strain) was constructed, injected into the adrenal gland of rats, and subsequently shown to transneuronally label the CNS autonomic neurons that project to the sympathoadrenal preganglionic neurons. Virally infected neurons were visualized with a one-step histochemical reaction using the Bluo-Gal substrate (halogenated indolyl-beta-D-galactoside) for the localization of beta-galactosidase activity. In some infected neurons, a Golgi-like staining of the primary and sometimes secondary dendrites could be obtained. For electron microscopic studies, the Bluo-Gal substrate produces an electron-dense reaction product that is easily identified at both low and high magnification. This virus may be useful for the study of the cell architecture and synaptic organization of transneuronally labeled neurons of functionally defined neural circuits. These results also demonstrate that it is possible to deliver foreign genes into specific chains of neurons in the mammalian CNS by means of the retrograde transneuronal vial labeling method.
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PMID:beta-Galactosidase expressing recombinant pseudorabies virus for light and electron microscopic study of transneuronally labeled CNS neurons. 165 2

We developed a sensitive two-site enzyme immunoassay (EIA) system for acidic fibroblast growth factor (aFGF), using a polyclonal antibody raised in rats. This assay is based on the sandwiching of the antigen between anti-aFGF antibody immunoglobulin G (IgG) coated on plates and biotinylated anti-aFGF antibody IgG; the detection of biotinylated IgG was performed by enzyme reaction of streptavidin-conjugated beta-D-galactosidase (beta-D-galactoside hydrolase; EC 3.2.1.23). Our system was specific for aFGF, because basic fibroblast growth factor, which shares a 55% homology of amino acid sequence with aFGF, hardly cross-reacted at all. The sensitivity of this system (0.2 ng/ml) enabled us to quantify endogenous immunoreactive aFGF in the CNS. Using this two-site EIA system, we examined the levels of aFGF in various regions of rat brain and their developmental changes. At the early stage of neonatal development, i.e., 2 days after birth, all brain regions registered low aFGF levels (less than 10 ng/g tissue). However, at the young adult stage (21- to 49-day-old animals), an extremely high level of aFGF (75-90 ng/g tissue) was found in the ponsmedulla; relatively high levels (30-40 ng/g tissue) were found in the diencephalon and mesencephalon; and comparatively low aFGF levels (5-15 ng/g tissue) were found in various other brain regions such as the frontal cortex, piriform cortex, hippocampus, olfactory bulb, cerebellum, and striatum. This marked change in the regional distribution of aFGF in the rat brain during postnatal development from 2 to 21 days after birth suggests that this factor plays a significant role in the brain during this period.
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PMID:Developmental changes in distribution of acidic fibroblast growth factor in rat brain evaluated by a sensitive two-site enzyme immunoassay. 170 23

We have developed a new single-step system, using a DEAE matrix, to immobilize and/or purify fusion proteins containing the choline-binding domain of the Streptococcus pneumoniae murein hydrolases. We have constructed a choline-binding-domain--beta-galactosidase chimera, which can be purified by this procedure and shows a high beta-galactosidase activity when immobilized in the column. A vector plasmid, pCUZ1, containing the lppp-5/lac promoter as well as 13 restriction sites, was constructed to facilitate the cloning and expression of gene fusions. This plasmid also allows the selection of recombinants by the well-known blue/white 5-bromo-4-chloro-3-indolyl-beta-D-galactoside procedure. A chimera between the choline-binding domain and the pneumococcal hemolysin was also constructed and purified using pCUZ1.
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PMID:Immobilization and single-step purification of fusion proteins using DEAE-cellulose. 173 Feb 20

We have devised a rapid method for examining the expression of a toxin gene following in vitro transfection using a bacterial beta-galactosidase (lacZ) gene as a reporter gene. Ricin A chain DNA and the lacZ gene, both under the control of the immunoglobulin gene promoter and enhancer, were transfected into mouse fibroblast cells (L cells). Transient expression of the lacZ gene was detected 2 days after transfection by histochemical staining of the transfectants with 5-bromo-3-indolyl-beta-D-galactoside. Cotransfection of the ricin A chain gene resulted in a progressive reduction in the number of lacZ transfectants as the expressed toxin killed the cells. A ricin construct with the intervening sequence from the human beta-actin gene required 4 days instead of 2 days to produce the toxic effect. This is a useful method for examining the expression of toxin gene in a cell.
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PMID:A method for detecting the expression of a toxic gene in cultured cells. 179 5


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