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:2.5.1.18 (glutathione S-transferase)
22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We previously purified a protein factor, named REKS (Ras-dependent Extracellular Signal-regulated Kinase (ERK)/mitogen-activated protein kinase Kinase (MEK) Stimulator), from Xenopus eggs by use of a cell-free assay system in which recombinant GTP gamma S (guanosine 5'-(3-O-thio)triphosphate)-Ki-Ras activates recombinant MEK. By use of this assay system, we purified here bovine REKS to near homogeneity from the cytosol fraction of bovine brain by successive chromatographies of Mono S, Mono Q, GTP gamma S-glutathione S-transferase-Ha-Ras-coupled glutathione-agarose, and Mono Q columns. It was composed of three proteins with masses of about 95, 32, and 30 kDa as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The 95-, 32-, and 30-kDa proteins were identified by immunoblot analysis to be B-Raf protein kinase, 14-3-3 protein, and 14-3-3 protein, respectively. Moreover, the REKS activity was specifically immunoprecipitated by an anti-B-Raf antibody. Bovine REKS was activated by lipid-modified GTP gamma S-Ki-Ras far more effectively than by a lipid-unmodified one. Lipid-modified GDP-Ki-Ras was inactive. Exogenous addition of 14-3-3 proteins stimulated further the REKS activity both in the presence and absence of GTP gamma S-Ki-Ras. These results indicate that at least one of the direct targets of Ras is B-Raf complexed with 14-3-3 proteins in bovine brain.
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PMID:Purification of a Ras-dependent mitogen-activated protein kinase kinase kinase from bovine brain cytosol and its identification as a complex of B-Raf and 14-3-3 proteins. 774 15

The 14-3-3 protein family has been implicated in growth factor signaling. We investigated whether 14-3-3 protein is involved in insulin signaling in 3T3L1 adipocytes. A significant amount of insulin receptor substrate 1 (IRS-1) was immunodetected in the immunoprecipitate with anti-14-3-3beta antibody at the basal condition. 100 nM insulin increased the amount of IRS-1 in the immunoprecipitate 2.5-fold. The effect of insulin was abolished by 100 nM wortmannin. An in vitro binding study revealed that glutathione S-transferase-14-3-3beta fusion protein directly associates with recombinant IRS-1. Pretreatment of recombinant IRS-1 with alkaline phosphatase clearly decreased this association. Because the recombinant IRS-1 was not phosphorylated on its tyrosine residues, the results suggest that serine/threonine phosphorylation of IRS-1 is responsible for the association. When the cells are treated with insulin, phosphatidylinositol 3'-kinase (PI3K) is supposed to complex either 14-3-3beta-IRS-1 or IRS-1. The 14-3-3beta-IRS-1-PI3K and IRS-1-PI3K complexes were separately prepared by a sequential immunoprecipitation, first with anti-14-3-3beta and then with anti-IRS-1 antibodies. The specific activity of the PI3K in the former was approximately half of that in the latter, suggesting that 14-3-3beta protein bound to IRS-1 inhibits insulin-stimulated lipid kinase activity of PI3K in 3T3L1 adipocytes.
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PMID:14-3-3beta protein associates with insulin receptor substrate 1 and decreases insulin-stimulated phosphatidylinositol 3'-kinase activity in 3T3L1 adipocytes. 942 53

We have previously reported on the cloning of the 14-3-3 protein of Schistosoma mansoni. Here, we evaluate the potential use of this protein as a vaccine candidate against infection by S. mansoni. Sm14-3-3 was expressed and purified either as a free protein or as a fusion protein to SjGST or MBP. Sera from mice infected with S. mansoni recognized both SjGST and 14-3-3, indicating that antibodies against these two proteins are induced in the course of the natural infection. Furthermore, mice immunized with either 14-3-3, GST or 14-3-3-GST, reacted with cercaria lysate. A cellular immune response was also detected, particularly in mice immunized with 14-3-3-GST. With respect to the effect on biological functions, antibodies to 14-3-3 and 14-3-3-GST caused 23-32% complement-mediated cytotoxcity of S. mansoni schistosomula compared to only 10-11% induced by either normal mouse serum, or GST alone. In challenge infection with S. mansoni, immunization with 14-3-3, either as a fusion protein or as a free protein, led to protection ranging from 25-46%, as determined by reduction of adult worm burden, while SjGST alone elicited only 0-8% protection and MBP alone did not elicit any protection.
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PMID:The 14-3-3 protein as a vaccine candidate against schistosomiasis. 1129 98

The diverse forms of protein phosphatase 1 (PP1) in vivo result from the association of the catalytic subunit with different regulatory subunits. We recently have described that PP1alpha is a Ras-activated Bad phosphatase that regulates IL-2 deprivation-induced apoptosis. With the yeast two-hybrid system, GST fusion proteins, indirect immunofluorescence, and coimmunoprecipitation, we found that Bcl-2 interacts with PP1alpha and Bad. In contrast, Bad did not interact with 14-3-3 protein. Bcl-2 depletion decreased phosphatase activity and association of PP1alpha to Bad. Bcl-2 contains the RIVAF motif, analogous to the well characterized R/KXV/IXF consensus motif shared by most PP1-interacting proteins. This sequence is involved in the binding of Bcl-2 to PP1alpha. Disruption of Bcl-2/PP1alpha association strongly decreased Bcl-2 and Bad-associated phosphatase activity and formation of the trimolecular complex. These results suggest that Bcl-2 targets PP1alpha to Bad.
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PMID:Bcl-2 targets protein phosphatase 1 alpha to Bad. 1139 Apr 85

The Arabidopsis ankyrin repeat-containing protein AKR2 was identified as a GF14(lambda)-interacting protein in a yeast two-hybrid screening (GF14(lambda) is a 14-3-3 protein). Reduced expression of AKR2 by using the antisense technique results in small necrotic areas in leaves accompanied by higher production of H2O2, similar to the hypersensitive response to pathogen infection in plant disease resistance. Transcripts of genes encoding pathogen-induced protein PR-1 (pathogen-related protein 1) and stress-responsive protein GST6 (glutathione S-transferase 6) are increased in antisense plants. The resistance to a bacterial pathogen infection was also increased by at least 10-fold in antisense plants. AKR2 also interacts with another GF14(lambda)-interacting protein, the ascorbate peroxidase 3 that scavenges H2O2 in plant cells. These data suggest that AKR2 may be a negative regulator of PR-1 expression, and is probably involved in the regulation of antioxidation metabolism that is shared by both disease resistance and stress responses.
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PMID:An ankyrin repeat-containing protein plays a role in both disease resistance and antioxidation metabolism. 1186 48

Protein kinase B/Akt (PKB/Akt) is a member of the ACG kinase family, which also includes protein kinase C, that phosphorylates a number of 14-3-3-binding proteins. 14-3-3 protein regulation of protein kinase C activity is modulated by 14-3-3 phosphorylation. We examined the hypothesis that PKB/Akt interacts with and phosphorylates 14-3-3zeta, leading to modulation of dimerization. By glutathione S-transferase pull-down, Akt precipitated recombinant 14-3-3zeta and endogenous 14-3-3zeta from HEK293 cell lysates. Recombinant active PKB/Akt phosphorylated recombinant 14-3-3zeta in an in vitro kinase assay. Transfection of active PKB/Akt into HEK293 cells resulted in phosphorylation of 14-3-3zeta. Based on a motif search of 14-3-3zeta, a potential PKB/Akt phosphorylation site, Ser-58, was mutated to alanine. PKB/Akt was unable to phosphorylate this mutant protein. Incubation of 14-3-3zeta with recombinant active PKB/Akt resulted in phosphorylation of 45% of the protein, as determined by a pI shift on two-dimensional electrophoresis, but 14-3-3zeta dimerization was not altered. These data indicate that PKB/Akt phosphorylates Ser-58 on 14-3-3zeta both in vitro and in intact cells. The functional relevance of this phosphorylation remains to be determined.
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PMID:Identification of 14-3-3zeta as a protein kinase B/Akt substrate. 1195 22

The 14-3-3 protein was shown to be present into the parasitophorous vacuole of Toxoplasma gondii-infected human monocyte cells and in the excreted/secreted antigens (ESA). The ESA 14-3-3 protein migrates electrophoretically as the cytosol and the main membranous 14-3-3 isoforms. The excretion/secretion of 14-3-3 was not sensitive to cycloheximide, a protein synthesis inhibitor, even at a concentration which inhibited the production of 14-3-3 inside the tachyzoites. Recombinant 14-3-3/GST protein was used to test the presence of 14-3-3 antibodies in different human sera. A positive immunoreactivity was observed with sera corresponding to acute toxoplasmosis and a possible involvement of 14-3-3 in host immunity is discussed.
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PMID:Characterization of an excreted/secreted antigen form of 14-3-3 protein in Toxoplasma gondii tachyzoites. 1510 15

Since drug resistance is a complex and multifactorial event involving activation/repression of multiple biochemical pathways, we used a proteomic approach to study cisplatin resistance and drug response in human tumor cell lines. The cervix squamous cell carcinoma cell line A431 and its cisplatin-resistant subline, A431/Pt, were used as a model system. The experimental set-up involved not just a two-way comparison of the control vs. the drug-resistant cell line, but also an acute cisplatin treatment of both cell lines, leading to a four-way comparison, as follows: 1) A431 vs. A431/Pt cells; 2) A431 vs. A431 cisplatin exposed cells; 3) A431/Pt vs. A431/Pt cisplatin exposed cells; 4) A431 cisplatin exposed cells vs. A431/Pt cisplatin exposed cells. We found modulation of proteins, which could be classified under various categories, such as molecular chaperones (e.g. heat-shock proteins HSP60, HSP90, HSC71, heat-shock cognate 71 kDa protein), Ca2+-binding proteins (e.g. calmodulin, calumenin), proteins involved in drug detoxification (such as peroxiredoxins PRX 2 and PRX 6, and glutathione-S-transferase, GST), anti-apoptotic proteins (such as 14-3-3 switched on in cisplatin-exposed cells) and ion channels (such as VDAC-1, voltage-dependent anion-selective channel). In particular, the basal levels of HSC71 and HSP60 were increased in A431/Pt cells as compared to A431 cells, and cisplatin exposure resulted in up-regulation of HSP60 and HSP90 only in A431 cells. Moreover, cisplatin exposure up-regulated the anti-apoptotic 14-3-3 protein in both cell lines, GST in sensitive cells and PRX6 in A431/Pt cells. These findings are consistent with a constitutive expression of defence factors by resistant cells and with activation by cisplatin of mechanisms acting to protect cells from drug-induced damage. This pattern of response, also observed in parental cells, could reflect an intrinsic resistance of this tumor type.
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PMID:A proteomic approach to cisplatin resistance in the cervix squamous cell carcinoma cell line A431. 1537 90

To search for the substrates of Ca2+/calmodulin-dependent protein kinase I (CaM-KI), we performed affinity chromatography purification using either the unphosphorylated or phosphorylated (at Thr177) GST-fused CaM-KI catalytic domain (residues 1-293, K49E) as the affinity ligand. Proteomic analysis was then carried out to identify the interacting proteins. In addition to the detection of two known CaM-KI substrates (CREB and synapsin I), we identified two Numb family proteins (Numb and Numbl) from rat tissues. These proteins were unphosphorylated and were bound only to the Thr177-phosphorylated CaM-KI catalytic domain. This finding is consistent with the results demonstrating that Numb and Numbl were efficiently and stoichiometrically phosphorylated in vitro at equivalent Ser residues (Ser264 in Numb and Ser304 in Numbl) by activated CaM-KI and also by two other CaM-Ks (CaM-KII and CaM-KIV). Using anti-phospho-Numb/Numbl antibody, we observed the phosphorylation of Numb family proteins in various rat tissue extracts, and we also detected the ionomycin-induced phosphorylation of endogenous Numb at Ser264 in COS-7 cells. The present results revealed that the Numb family proteins are phosphorylated in vivo as well as in vitro. Furthermore, we found that the recruitment of 14-3-3 proteins was the functional consequence of the phosphorylation of the Numb family proteins. Interaction of 14-3-3 protein with phosphorylated Numbl-blocked dephosphorylation of Ser304. Taken together, these results indicate that the Numb family proteins may be intracellular targets for CaM-Ks, and they may also be regulated by phosphorylation-dependent interaction with 14-3-3 protein.
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PMID:Phosphorylation of Numb family proteins. Possible involvement of Ca2+/calmodulin-dependent protein kinases. 1610 44

Fusarium head blight (FHB) caused by Fusarium graminearum is a destructive disease of wheat and barley. It causes economic losses due to reduction in both yield and quality. Although FHB resistance has been well documented and resistant cultivars have been developed to reduce incidence and severity of FHB, there is a limited understanding of the molecular mechanisms involved in plant resistance against the infection and spread of F. graminearum. In the current study, 2-dimensional displays of proteins extracted from wheat spikelets infected with F. graminearum were compared with those from spikelets inoculated with sterile H2O. Fifteen protein spots were detected that were either induced (qualitatively different) or upregulated (quantitatively increased) following F. graminearum infection of spikelets of 'Ning7840', a resistant cultivar. These proteins were identified by LC-MS/MS analysis. Proteins with an antioxidant function such as superoxide dismutase, dehydroascorbate reductase, and glutathione S-transferases (GSTs) were upregulated or induced 5 d after inoculation with F. graminearum, indicating an oxidative burst of H2O2 inside the tissues infected by FHB. An ascorbate-glutathione cycle is likely involved in reduction of H2O2. Expression of proteins with highest similarity to dehydroascorbate reductase and TaGSTF5 (a glutathione S-transferase) differed following FHB infection in susceptible and resistant cultivars. A 14-3-3 protein homolog was also upregulated in FHB-infected spikelets. In addition, a PR-2 protein (beta-1, 3 glucanase) was upregulated in FHB-infected spikes, which is in accord with a previous study that analyzed transcript accumulation.
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PMID:Identification of proteins induced or upregulated by Fusarium head blight infection in the spikes of hexaploid wheat (Triticum aestivum). 1639 83


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