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)

Molecular analysis of the human beta-galactosidase gene revealed six different mutations in 10 of 11 Japanese GM1-gangliosidosis patients. They were the only abnormalities in each allele examined in this study. A 165-nucleotide duplication (positions 1103-1267) was found in two infantile patients, producing an abnormally large mRNA; one patient was probably a homozygote, and the other was a heterozygote of this mutation. The other two infantile patients had different mutations; a 123 Gly(GGG)----Arg(AGG) mutation in one patient and a 316 Tyr(TAT)----Cys(TGT) mutation in the other. A 201 Arg(CGC)----Cys(TGC) mutation, eliminating a BspMI site, was detected in a late-infantile/juvenile patient; the restriction-site analysis of amplified genomic DNA confirmed his heterozygosity for this mutation. A 51 Ile(ATC)----Thr(ACC) mutation was found in all five adult/chronic patients examined in this study. It created a SauI site, and restriction-site analysis confirmed that four patients were homozygous mutants. The other was a compound heterozygote for this mutation and another 457 Arg(CGA)----Gln(CAA) mutation. These mutant genes expressed markedly decreased or completely deficient enzyme activities in beta-galactosidase-deficient human fibroblasts transformed by adenovirus-SV40 recombinants. We conclude that gene mutations are heterogeneous in GM1-gangliosidosis but that the 51 Ile(ATC)----Thr(ACC) mutation is common among the Japanese adult/chronic cases. Genotype-phenotype correlations in GM1-gangliosidosis are briefly discussed.
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PMID:Human beta-galactosidase gene mutations in GM1-gangliosidosis: a common mutation among Japanese adult/chronic cases. 190

In this study we have reconstituted transactivation of gene expression by the human glucocorticoid receptor in the yeast, Saccharomyces cerevisiae. We have expressed the C-terminal half of the human glucocorticoid receptor (residues 415-777), the smallest derivative that can be expected to function as a ligand-dependent activator of transcription, in yeast cells. The function of the expressed protein has been assayed using a reporter gene consisting of the beta-galactosidase gene from Escherichia coli fused to the yeast iso-1-cytochrome c promoter with a glucocorticoid-responsive element from the rat tyrosine aminotransferase gene upstream. Transactivation of expression from the reporter gene by the expressed receptor is seen only in the presence of steroid hormones with glucocorticoid activity and occurs via specific interaction of receptor with the glucocorticoid-responsive element upstream of the reporter gene. This result is different from those obtained for the estrogen receptor in which a similar derivative was not functional in yeast. This suggests that the well documented conservation of structure and function between steroid receptors may not extend to the transactivation domains. Our results also suggest that the mechanism by which receptors are sequestered in an inactive, non-DNA binding state in the absence of ligand may be functionally conserved in yeast. In support of this we show evidence that the expressed receptor is associated with the yeast molecular weight 90,000 heat shock protein as seen in mammalian cells.
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PMID:Ligand-specific transactivation of gene expression by a derivative of the human glucocorticoid receptor expressed in yeast. 220 60

A gene encoding the human immunodeficiency virus-1 (HIV-1) TAT protein was chemically synthesized and expressed in HeLa cells and in a cell-free system. To facilitate both the assembly of the synthetic gene and further mutagenesis and gene fusion studies, several unique restriction endonuclease cleavage sites were included in the coding sequence without altering the encoded protein sequence. The synthetic TAT coding sequence was fused to a translation start signal and placed under SV40 early transcriptional control. Co-transfection of the TAT-encoding synthetic gene together with a reporter gene (chloramphenical acetyl transferase or beta-galactosidase) linked to an HIV LTR confirmed that the synthetic gene product exhibits similar activity to TAT expressed from HIV genomic DNA in the transactivation of the LTR. TAT mRNA prepared by cell-free transcription of the synthetic TAT coding sequence was also shown to produce functional TAT following microinjection into HeLa-derived cells containing an integrated reporter gene with the HIV LTR linked to beta-galactosidase.
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PMID:Chemical synthesis and expression of a gene encoding HIV-1 TAT protein. 254

Adrenalectomy of suckling rats is complicated by a high mortality rate, caused by the loss of blood (early mortality) and by the disturbed sodium-potassium balance (late mortality). Treatment of the abdominal cavity with a thrombin solution and a daily administration of deoxycorticosterone glucoside (DOC) decrease the total mortality remarkably. DOC treatment has no influence on renal beta-glucosidase and beta-galactosidase as well as on hepatic tyrosine aminotransferase activity, whereas hepatic serine dehydratase activity exhibits a time- and dosage-dependent response to this hormone. The DOC effect is very likely a consequence of the glucocorticoid-like action of the synthetic hormone, which competes with the endogenous glucocorticoids for the hepatic receptor molecules.
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PMID:An improved method for adrenalectomy of suckling rats. The influence of thrombin treatment and deoxycorticosterone substitution on survival and on hepatic and renal enzyme activities. 287 58

From a review of past studies and the report of new studies from our laboratory, this article provides strong evidence to show that WB-F344 (WB) rat liver epithelial cells are stem-like precursor cells for hepatocytes. WB cells are structurally and phenotypically simple epithelial cells that were isolated from the liver of an adult male Fischer 344 rat, under conditions that excluded their origin from hepatocytes in vivo. WB cells express a phenotypic repertory that overlaps, but is distinct from, that of both hepatocytes and bile duct epithelial cells. The complex phenotype of WB cells is compatible with their being embryonic or undifferentiated variants of either hepatocytes or bile duct epithelial cells. When WB cells are tagged genetically with genes for bacterial beta-galactosidase and neomycin resistance (BAG2-WB), they and their progeny can be distinguished from parental WB cells and hepatocytes by the expression of these gene products. Progeny of BAG2-WB cells that were transplanted into the liver parenchyma of syngeneic rats integrated into hepatic plates and acquired the morphological and functional attributes of adjacent host hepatocytes; the progeny of BAG2-WB cells in the liver express albumin, tyrosine aminotransferase, alpha-1-antitrypsin, and transferrin. We also demonstrate that progeny of BAG2-WB cells can be recovered from livers into which they have been transplanted, which may allow the elucidation of alterations in gene expression that accompany their differentiation.
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PMID:Isolation, culture, and transplantation of rat hepatocytic precursor (stem-like) cells. 823 70

Protein transduction domains (PTDs), such as the third helix of the Drosophila Antennapedia homeobox gene (Antp) and the HIV TAT PTD, possess a characteristic positive charge on the basis of their enrichment for arginine and lysine residues. To determine whether cationic peptides are able to function as protein transduction domains, 12-mer peptide sequences from an M13 phage library were selected for synthesis on the basis of their varying cationic charge content. In addition, polylysine and polyarginine peptides were synthesized in order to assess the effect of charge contribution in protein transduction. Coupling of the biotinylated peptides to avidin-beta-galactosidase facilitated transduction in a wide variety of cell lines and primary cells, including islet beta-cells, synovial cells, polarized airway epithelial cells, dendritic cells, myoblasts, and tumor cells. Two of the peptides, PTD-4 and PTD-5, mediated transduction nearly 600-fold more efficiently than a random control peptide, but with an efficiency similar to the TAT PTD and the 12 mers of polylysine and polyarginine. Furthermore, confocal analysis of biotinylated peptide-streptavidin-Cy3 conjugates demonstrated that the internalized PTDs are found in both the nuclei and the cytoplasm of treated cells. When tested in vivo, the PTDs were able to facilitate efficient and rapid protein delivery into rabbit synovium and mouse solid tumors following intraarticular and intratumoral administration, respectively. These novel PTDs can be used to transfer therapeutic proteins and DNA for the treatment of a wide variety of diseases, including arthritis and cancer.
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PMID:Characterization of a class of cationic peptides able to facilitate efficient protein transduction in vitro and in vivo. 1102 Mar 49

We have studied the transduction of TAT-HA-beta-galactosidase fusion protein into two cell lines of rat salivary gland origin, A5 and C6-21, into cells of fetal mouse submandibular glands in organ culture, and into rat submandibular gland after retrograde duct injection, using a histochemical method to demonstrate beta-galactosidase activity. Transduction of the fusion protein into A5 and C6-21 cells was concentration- and time-dependent. Therefore, the intensity of the beta-galactosidase staining, which was cytoplasmic, was less after 1 hr of exposure compared to exposures up to 24 hr. However, the fusion protein was transduced into 100% of both types of cultured cells. When explants of mouse fetuses at 13 days of gestation were exposed to the fusion proteins, both epithelial and mesenchymal cells were stained for the enzyme, with a conspicuous accumulation of the reaction product at perinuclear cytoplasmic regions. The histochemical staining of the mesenchymal cells was more intense compared to that seen in epithelial cells. TAT-HA-beta-galactosidase fusion protein was also delivered to rat submandibular glands by retrograde duct injection. Histochemical staining for beta-galactosidase activity of cryostat sections prepared from the injected glands revealed that the transduction of the fusion protein was also time- and dose-dependent. In the glands of rats sacrificed from 10 min to 1 hr after the retrograde injection, essentially all acinar and duct cells showed cytoplasmic staining. The intensity of the staining then declined, and was not seen in the glands of rats killed 24 hr after the injection of the fusion proteins. These results indicate that a full-length, active TAT fusion protein can be targeted to salivary gland cells both in vitro and in vivo to analyze physiological, developmental, and pathophysiological processes.
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PMID:Transduction of TAT-HA-beta-galactosidase fusion protein into salivary gland-derived cells and organ cultures of the developing gland, and into rat submandibular gland in vivo. 1103 88

The resounding success of a new immunosuppressive regimen known as the Edmonton protocol demonstrates that islet cell transplantation is becoming a therapeutic reality for diabetes. However, under the Edmonton protocol, a single donor does not provide enough islets to attain the insulin independence of a transplant recipient. This limitation is mainly caused by islet apoptosis triggered during isolation. In this study, we describe a highly efficient system of transiently transferring anti-apoptotic proteins into pancreatic islets, thus opening an exciting new therapeutic opportunity to improve the viability of transplantable islets. We fused beta-galactosidase to the 11-amino acid residues that constitute the protein transduction domain (PTD) of the HIV/TAT protein and transduced pancreatic islets ex vivo with this fusion protein in a dose-dependent manner with >80% efficiency. We observed that transduction of the anti-apoptotic proteins Bcl-X(L) and PEA-15 fused to TAT/PTD prevented apoptosis induced by tumor necrosis factor-alpha in a pancreatic beta-cell line, indicating that TAT/PTD anti-apoptotic proteins retained their biological activity. Finally, we demonstrated that TAT-fusion proteins did not affect the insulin secretion capability of islets, as determined by glucose static incubation and by reversion of hyperglycemia in diabetic immunodeficient mice.
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PMID:Proteins linked to a protein transduction domain efficiently transduce pancreatic islets. 1147 28

Human solid tumors contain hypoxic regions that have considerably lower oxygen tension than normal tissues. These impart resistance to radiotherapy and anticancer chemotherapy, as well as predisposing to increased tumor metastases. To develop a potentially therapeutic protein drug highly specific for solid tumors, we constructed fusion proteins selectively stabilized in hypoxic tumor cells. A model fusion protein, oxygen-dependent degradation (ODD)-beta-galactosidase (beta-Gal), composed of a part of the ODD domain of hypoxia-inducible factor-1alpha fused to beta-Gal, showed increased stability in cultured cells under a hypoxia-mimic condition. When ODD-beta-Gal was further fused to the HIV-TAT protein transduction domain (TAT(47-57)) and i.p. injected to a tumor-bearing mouse, the biologically active fusion protein was specifically stabilized in solid tumors but was hardly detected in the normal tissue. Furthermore, when wild-type (WT) caspase-3 (Casp3(WT)) or its catalytically inactive mutant was fused to TAT-ODD and i.p. injected to a tumor-bearing mouse, the size of tumors was reduced by the administration of TAT-ODD-Casp3(WT) but not by TAT-ODD-mutant Casp3. TAT-ODD-Casp3(WT) did not cause any obvious side effects on tumor-bearing mice, suggesting specific stabilization and activation of the fusion protein in the hypoxic tumor cells. These results suggest that the combination of protein therapy using a cytotoxic TAT-ODD fusion protein with radiotherapy and chemotherapy may provide a new strategy for annihilating solid tumors.
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PMID:Antitumor effect of TAT-oxygen-dependent degradation-caspase-3 fusion protein specifically stabilized and activated in hypoxic tumor cells. 1192 18

The delivery of proteins across the blood-brain barrier is severely limited by the proteins' size and biochemical properties. Eleven-amino acid human immunodeficiency virus TAT protein is able to cross cell membranes even when coupled with larger peptides. We evaluated whether TAT-Bcl-X(L) fusion protein is protective in focal ischemia. Mice underwent 30 or 90 minutes of intraluminal middle cerebral artery thread occlusion. TAT-Bcl-X(L), TAT-beta-galactosidase, or TAT-GFP (0.6 nmol each) were applied intravenously over 10 minutes either 1 hour before or immediately after ischemia. Additional animals received no TAT protein infusions. We show that the brain tissue is progressively transduced with TAT proteins within 3 to 4 hours after intravenous delivery. We provide evidence that TAT-Bcl-X(L) treatment reduces infarct volume and neurological deficits after long ischemic insults lasting 90 minutes, when applied both before and after ischemia. After short insults, lasting only 30 minutes, TAT-Bcl-X(L) further diminishes the number of caspase-3-reactive and DNA fragmented cells and increases the number of viable neurons in the striatum. Our results indicate that TAT fusion proteins are elegant and powerful tools that might be of clinical interest for stroke treatment, because factors may be intravenously applied. Thus, fusion proteins may open fascinating perspectives for future research.
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PMID:Intravenous TAT-Bcl-Xl is protective after middle cerebral artery occlusion in mice. 1240 59


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