Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.5.1.18 (glutathione S-transferase)
 22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Olefinic compounds are commercially valuable because they form useful polymeric substances. The same chemical property (presence of double bonds) that makes the olefins useful may also cause them to be toxic in the body. The double bonds of olefins can be oxidized by cytochrome P450 enzymes to epoxides, which are electrophiles that can react with DNA and may cause alterations in the genetic information carried by that macromolecule. Epoxides can be rendered inactive toward DNA by binding to proteins, by hydrolysis to diols through epoxide hydrolase enzymes (EHs), or by forming conjugates with glutathione via glutathione S-transferase (GST) activities. The balance between the oxidizing enzymatic activities and the hydrolyzing or conjugating enzymatic activities in the livers of different species can influence the potential toxicity of the olefins. The location of the enzymes and the potential for concerted reactions in which epoxides are inactivated immediately after formation will also influence the potential toxicity of the olefins. Cytochrome P450 enzymes and EHs are in microsomes located in the rough endoplasmic reticulum surrounding the nucleus where the DNA is located. GST is in the cytoplasm of the cell. In the case of 1,3-butadiene (BD), such enzymatic differences may strongly influence the toxicity in different species. The mouse, in which BD is a potent multi-site carcinogen, has the lowest microsomal EH activity of any species. This allows the monoepoxides formed in the microsomes by cytochrome P450 enzymes to be further oxidized to the highly genotoxic diepoxide (DEB), and both epoxides can either be released into the blood for distribution throughout the body or can react with DNA in the nucleus. The rat, in which BD is a weak carcinogen, has much higher levels of microsomal EH, and only trace amounts of DEB enter the bloodstream. Major BD metabolites in primates suggest that the hydrolysis pathway is even more prominent in primates than in rats. Data suggest that BD will be much less toxic in primates than in mice. Considering these quantitative differences in metabolism may help to reduce the uncertainties in extrapolating animal data on olefin toxicity to health risk assessments for humans exposed to the compounds.
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PMID:Species differences in the metabolism of olefins: implications for risk assessment. 1139 81

Results from our previous study suggest that cyclooxygenase-2 (COX-2) induced by phorbol 12-myristate 13-acetate (PMA) may be localized to caveolae-like structures (Liou, J.-Y., Shyue, S.-K., Tsai, M.-J., Chung, C.-L., Chu, K.-Y., and Wu, K. K. (2000) J. Biol. Chem. 275, 15314-15320). In this study, we determined subcellular localization of COX-2 and caveolin-1 by confocal microscopy. COX-2 in human foreskin fibroblasts stimulated by PMA (100 nm) or interleukin-1beta (1 ng/ml) for 6 h was localized to plasma membrane in addition to endoplasmic reticulum and nuclear envelope. Caveolin-1 was localized to plasma membrane, and image overlay showed colocalization of COX-2 with caveolin-1. This was confirmed by the presence of COX-2 and caveolin-1 in the detergent-insoluble membrane fraction of cells stimulated by PMA. Immunoprecipitation showed complex formation of COX-2 with caveolin-1 in a time-dependent manner. A larger quantity of COX-2 was complexed with caveolin-1 in PMA-treated than in interleukin-1beta-treated cells. Purified COX-2 complexed with glutathione S-transferase-fused caveolin-1, which was not inhibited by the scaffolding domain peptide. Caveolin-1-bound COX-2 was catalytically active, and its activity was not inhibited by the scaffolding domain peptide. These results suggest that COX-2 induced by PMA and interleukin-1beta is colocalized with caveolin-1 in the segregated caveolae compartment. Because caveolae are rich in signaling molecules, this COX-2 compartment may play an important role in diverse pathophysiological processes.
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PMID:Colocalization and interaction of cyclooxygenase-2 with caveolin-1 in human fibroblasts. 1143 74

Although all mammalian COPII components have now been cloned, little is known of their interactions with other regulatory proteins involved in exit from the endoplasmic reticulum (ER). We report here that a mammalian protein (Yip1A) that is about 31% identical to S. cerevisiae and which interacts with and modulates COPII-mediated ER-Golgi transport. Yip1A transcripts are ubiquitously expressed. Transcripts of a related mammalian homologue, Yip1B, are found specifically in the heart. Indirect immunofluorescence microscopy revealed that Yip1A is localized to vesicular structures that are concentrated at the perinuclear region. The structures marked by Yip1A co-localized with Sec31A and Sec13, components of the COPII coat protein complex. Immunoelectron microscopy also showed that Yip1A co-localizes with Sec13 at ER exit sites. Overexpression of the hydrophilic N terminus of Yip1A arrests ER-Golgi transport of the vesicular stomatitis G protein and causes fragmentation and dispersion of the Golgi apparatus. A glutathione S-transferase fusion protein with the hydrophilic N terminus of Yip1A (GST-Yip1A) is able to bind to and deplete vital components from rat liver cytosol that is essential for in vitro vesicular stomatitis G transport. Peptide sequence analysis of cytosolic proteins that are specifically bound to GST-Yip1A revealed, among other proteins, mammalian COPII components Sec23 and Sec24. A highly conserved domain at the N terminus of Yip1A is required for Sec23/Sec24 interaction. Our results suggest that Yip1A is involved in the regulation of ER-Golgi traffic at the level of ER exit sites.
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PMID:A membrane protein enriched in endoplasmic reticulum exit sites interacts with COPII. 1148 4

Fusion constructs of partial sequences of triadin that contain green fluorescent protein at the N-terminus and glutathione transferase at the C-terminus have been expressed in human embryonic kidney -293 cells. A comparison of the subcellular disposition of a range of triadin fusion peptides indicates localization either to a few large organelles as a default target or to endoplasmic reticulum when amino acids 68-98 are present and structurally intact. Fluorescence from the conjugate of monochlorobimane with glutathione identifies whether the C-terminus has a cytoplasmic or luminal location. A stable transit of the membrane occurs in triadin2-98. Triadin2-117 and 2-267 give both cytoplasmic and luminal C-termini. Both triadin89-117 and triadin89-267 distribute in membranes, but do not cross them. The data are interpreted to indicate that cardiac triadin contains an alpha-helical membrane transit through the hydrophobic domain, 49-68, and a membrane association through the short hydrophobic domain, 102-114.
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PMID:Membrane topography of cardiac triadin. 1181 49

Acyl-coenzyme A:cholesterol acyltransferase (ACAT) is an enzyme involved in cellular cholesterol homeostasis and atherosclerosis. ACAT1 is an allosteric enzyme responding to its substrate cholesterol in a sigmoidal manner. It is a homotetrameric protein that spans the membrane multiple times, with its N-terminal 131 hydrophilic amino acids residing at the cytoplasmic side of the endoplasmic reticulum. This region contains two closely linked putative alpha-helices. Our current studies show that this region contains a dimer-forming motif. Adding this motif to the bacterial glutathione S-transferase (GST) converted the homodimeric GST to a tetrameric fusion protein. Conversely, deleting this motif from the full-length ACAT1 converted the enzyme from a homotetramer to a homodimer. The dimeric ACAT1 remains enzymatically active. Its biochemical characteristics, including the sigmoidal response to cholesterol, the IC(50) value toward a specific ACAT inhibitor, and sensitivity toward heat inactivation, are essentially unaltered. On the other hand, the dimeric ACAT1 exhibits a 5-10-fold increase in the V(max) of the overall reaction and a 2.2-fold increase in the K(m) for oleoyl-coenzyme. Thus, deleting the dimer-forming motif near the N-terminus changes ACAT1 from its tetrameric form to a dimeric form and increases its catalytic efficiency.
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PMID:Role of the N-terminal hydrophilic domain of acyl-coenzyme A:cholesterol acyltransferase 1 on the enzyme's quaternary structure and catalytic efficiency. 1188 94

Ptd(4,5)P(2) is thought to promote and organize a wide range of cellular functions, including vesicular membrane traffic and cytoskeletal dynamics, by recruiting functional protein complexes to restricted locations in cellular membranes. However, little is known about the distribution of PtdIns(4,5)P(2) in the cell at high resolution. We have used the pleckstrin homology (PH) domain of phospholipase delta(1) (PLCdelta(1)), narrowly specific for PtdIns(4,5)P(2), to map the distribution of the lipid in astrocytoma and A431 cells. We applied the glutathione S-transferase-tagged PLCdelta(1) PH domain (PLCdelta(1)PH-GST) in an on-section labelling approach which avoids transfection procedures. Here we demonstrate PtdIns(4,5)P(2) labelling in the plasma membrane, and also in intracellular membranes, including Golgi (mainly stack), endosomes and endoplasmic reticulum, as well as in electron-dense structures within the nucleus. At the plasma membrane, labelling was more concentrated over lamellipodia, but not in caveolae, which contained less than 10% of the total cell-surface labelling. A dramatic decrease in signal over labelled compartments was observed on preincubation with the cognate headgroup [Ins(1,4,5)P(3)], and plasma-membrane labelling was substantially decreased after stimulation with thrombin-receptor-activating peptide (SFLLRN in the one-letter amino acid code), a treatment which markedly diminishes PtdIns(4,5)P(2) levels. Thus we have developed a highly selective method for mapping the PtdIns(4,5)P(2) distribution within cells at high resolution, and our data provide direct evidence for this lipid at key functional locations.
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PMID:Subcellular localization of phosphatidylinositol 4,5-bisphosphate using the pleckstrin homology domain of phospholipase C delta1. 1196 66

We previously reported the identification of a novel gene, Bdm1/NDRG4, that was expressed predominantly in the postnatal rat brain and might possibly play a role in this process. We describe here the characterization of a NDRG4 protein in a developing and maturing rat brain. Antibody raised against glutathione S-transferase (GST)-NDRG4 fusion protein recognized four protein species of 38, 39, 41, and 45 kDa on Western blotting of proteins from differently staged rat brains. The 38-kDa form was revealed after birth, and the amount of this species peaked on postnatal day 15. The 39-kDa form became detectable after postnatal week 6. The 41-kDa form appeared late in embryogenesis, increased by postnatal day 15, and disappeared at postnatal week 6. The 45-kDa form was abundant during the late embryonic period and slightly decreased after birth. Subcellular fractionation of cerebra indicated that the NDRG4 protein was distributed mainly in the mitochondria and endoplasmic reticulum (ER). Detergent solubility assays and protease susceptibility demonstrated that in the ER NDRG4 protein is membrane-associated and luminally oriented. The 45-kDa isoform was induced during NGF-mediated neuronal differentiation of PC12 cells, but not by tunicamycin which causes ER stress. Differential expressions of NDRG4 protein isoforms may be a mechanism for modifying the NDRG4 function and for the formation of a functioning nervous system.
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PMID:Molecular characterization of NDRG4/Bdm1 protein isoforms that are differentially regulated during rat brain development. 1197 92

Oxysterol-binding protein (OSBP) is 1 of 12 related proteins implicated in the regulation of vesicle transport and sterol homeostasis. A yeast two-hybrid screen using full-length OSBP as bait was undertaken to identify partner proteins that would provide clues to the function of OSBP. This resulted in the cloning of vesicle-associated membrane protein-associated protein-A (VAP-A), a syntaxin-like protein implicated in endoplasmic reticulum (ER)/Golgi vesicle transport, and phospholipid regulation in mammalian cells and yeast, respectively. By using a combination of yeast two-hybrid, glutathione S-transferase pull-down and immunoprecipitation experiments, the VAP-A-binding region in OSBP was localized to amino acids 351-442. This region did not include the pleckstrin homology (PH) domain but overlapped with the N terminus of the oxysterol binding and OSBP homology domains. C- and N-terminal truncations or deletions of VAP prevented interaction with OSBP but did not affect VAP multimerization. Although the OSBP PH domain was not necessary for VAP-A binding in vitro, interaction with VAP-A was enhanced in cells by mutation of the conserved PH domain tryptophan (OSBP W174A) or deletion of the C-terminal half of the PH domain (OSBP Delta 132-182). OSBP W174A retained oxysterol binding activity, association with phospholipid vesicles via the PH domain, and localized with VAP in unusual ER-associated structures. At 40 degrees C, misfolded ts045-vesicular stomatitis virus G protein fused to green fluorescent protein was co-localized with VAP-A/OSBP W174A structures on the ER but was exported to the Golgi when folded normally at 32 degrees C. A fluorescent ceramide analogue also accumulated in these ER inclusions, and export to the Golgi was partially inhibited as indicated by decreased Golgi staining and a 30% reduction in sphingomyelin synthesis. These studies show that OSBP binding to the ER and Golgi apparatus is regulated by its PH domain and VAP interactions, and the complex is involved at a stage of protein and ceramide transport from the ER.
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PMID:Vesicle-associated membrane protein-associated protein-A (VAP-A) interacts with the oxysterol-binding protein to modify export from the endoplasmic reticulum. 1202 75

VHL is part of an SCF related E3-ubiquitin ligase complex with 'gatekeeper' function in renal carcinoma. However, no mutations have been identified in VHL interacting proteins in wild type VHL tumors. We previously reported that the TRC8 gene was interrupted by a t(3;8) translocation in a family with hereditary renal and non-medullary thyroid cancer. TRC8 encodes a multi-membrane spanning protein containing a RING-H2 finger with in vitro ubiquitin ligase activity. We isolated the Drosophila homologue, DTrc8, and studied its function by genetic manipulations and a yeast 2-hybrid screen. Human and Drosophila TRC8 proteins localize to the endoplasmic reticulum. Loss of either DTrc8 or DVhl resulted in an identical ventral midline defect. Direct interaction between DTrc8 and DVhl was confirmed by GST-pulldown and co-immunoprecipitation experiments. CSN-5/JAB1 is a component of the COP9 signalosome, recently shown to regulate SCF function. We found that DTrc8 physically interacts with CSN-5 and that human JAB1 localization is dependent on VHL mutant status. Lastly, overexpression of DTrc8 inhibited growth consistent with its presumed role as a tumor suppressor gene. Thus, VHL, TRC8, and JAB1 appear to be linked both physically and functionally and all three may participate in the development of kidney cancer.
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PMID:The TRC8 hereditary kidney cancer gene suppresses growth and functions with VHL in a common pathway. 1203 52

The full-length cDNA clone of a novel GRP78-binding protein (GBP) was isolated from rat brain using PCR-selected cDNA subtraction. GBP was predominantly expressed in neuronal cells among various brain tissues. GBP mRNA was already detected in the E12 brain and then gradually increased to reach a peak within P0-2 weeks after birth. GBP expression in the brain decreased age-dependently to approximately 30% of the postnatal level at 12 months. GBP encoded 1021 amino acids and was predicted to have two transmembrane regions and glutamic acid- and proline-rich regions. Because the sequence of GBP offered few clues to the possible function, we performed a GST-tagged GBP pull-down assay in PC12 lysates and identified GRP78, one of the heat shock proteins, as a counterpart. Observation of COS7 cells expressing green fluorescent protein- or Myc-tagged GBP showed that GBP was localized in the endoplasmic reticulum-Golgi domain where BODIPY 558/568 (4,4-difluro-5-(2-thienyl)-4-bora-3alpha,4alpha-diaza-S-indacene)-labeled brefeldin A accumulated. To investigate a biological role for GBP, we established Neuro2a cells stably expressing Myc-tagged GBP. Overexpression of GBP did not affect cell growth or morphological features but attenuated the time-dependent decrease in cell viability caused by serum deprivation compared with control cells. After 48 h of serum starvation, Neuro2a cells overexpressing GBP were resistant to the cell death induced by serum withdrawal. These results suggest that GBP would have a relevant functional role in embryonic and postnatal development of the brain.
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PMID:Cloning and characterization of a novel GRP78-binding protein in the rat brain. 1251 90


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