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)

Tyrosine phosphorylation of cellular proteins is an early and an essential step in T cell receptor-mediated lymphocyte activation. Tyrosine phosphorylation of transmembrane receptor chains (such as zeta and CD3 chains) and membrane-associated proteins provides docking sites for SH2 domains of adaptor proteins and signaling enzymes, resulting in their recruitment in the vicinity of activated receptors. pp36/38 is a prominent substrate of early tyrosine phosphorylation upon stimulation through the T cell receptor. The tyrosine-phosphorylated form of pp36/38 is membrane-associated and directly interacts with phospholipase C-gamma 1 and Grb2, providing one mechanism to recruit downstream effectors to the cell membrane. Here, we demonstrate that in Jurkat T cells, pp36/38 associates with the p85 subunit of phosphatidylinositol 3-kinase (PI-3-K p85) in an activation-dependent manner. Association of pp36/38 with PI-3-K p85 was confirmed by transfection of a hemagglutinin-tagged p85 alpha cDNA into Jurkat cells followed by anti-hemagglutinin immunoprecipitation. In vitro binding experiments with glutathione S-transferase fusion proteins of PI-3-K p85 demonstrated that the SH2 domains, but not the SH3 domain, mediated binding to pp36/38. This binding was selectively abrogated by phosphopeptides that bind to p85 SH2 domains with high affinity. Filter binding assays demonstrated that association between pp36/38 and PI-3-K p85 SH2 domains was due to direct binding. These results strongly suggest the role of pp36/38 in recruiting PI-3-K to the cell membrane and further support the idea that pp36/38 is a multifunctional docking protein for SH2 domain-containing signaling proteins in T cells.
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PMID:T cell activation-dependent association between the p85 subunit of the phosphatidylinositol 3-kinase and Grb2/phospholipase C-gamma 1-binding phosphotyrosyl protein pp36/38. 754 53

The angiotensin II type-1 (AT1) receptor, a G protein-coupled receptor, lacks intrinsic kinase activity. However, recent data show that angiotensin II (Ang II) stimulates tyrosine phosphorylation of phospholipase C-gamma 1 (PLC-gamma 1), Stat91 (one of the signal transducers and activators of transcription), and paxillin in vascular smooth muscle cells. The tyrosine kinases responsible for these phosphorylation events are unknown. Src family kinases have been shown to phosphorylate PLC-gamma 1 and to be activated by G protein-coupled receptors. We hypothesized that pp60c-src associates with the AT1 receptor and is activated after Ang II stimulation of smooth muscle cells. We immunoprecipitated pp60c-src from Ang II-stimulated vascular smooth muscle cells and measured pp60c-src activity by autophosphorylation and by phosphorylation of enolase. Both assays demonstrated an approximately threefold increase in pp60c-src activity within 1 minute. A similar increase in Ang II-stimulated pp60c-src activity was observed in Chinese hamster ovary cells transfected with the AT1 receptor but not in untransfected cells. These data are the first to show that pp60c-src is activated by Ang II. To determine if pp60c-src associated with the AT1 receptor, the AT1 receptor was immunoprecipitated (with two different antibodies), and Western blots were performed with two different anti-pp60c-src antibodies. No pp60c-src was detected. In addition, direct interaction between the AT1 receptor and pp60c-src could not be demonstrated by using a glutathione S-transferase (GST)-AT1 fusion protein to bind proteins from cell lysates stimulated by Ang II.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Angiotensin II activates pp60c-src in vascular smooth muscle cells. 758 16

The activation of protein tyrosine kinases is a critical event in T cell antigen receptor (TCR)-mediated signaling. One substrate of the TCR-activated protein tyrosine kinase pathway is a 76-kDa protein (pp76) that associates with the adaptor protein Grb2. In this report we describe the purification of pp76 and the molecular cloning of its cDNA, which encodes a novel 533-amino acid protein with a single carboxyl-terminal Src homology 2 (SH2) domain. Although no recognizable motifs related to tyrosine, serine/threonine, or lipid kinase domains are present in the predicted amino acid sequence, it contains several potential motifs recognized by SH2 and SH3 domains. A cDNA encoding the murine homologue of pp76 was also isolated and predicts a protein with 84% amino acid identity to human pp76. Northern analysis demonstrates that pp76 mRNA is expressed solely in peripheral blood leukocytes, thymus, and spleen; and in human T cell, B cell and monocytic cell lines. In vitro translation of pp76 cDNA gives rise to a single product of 76 kDa that associates with a GST/Grb2 fusion protein, demonstrating a direct association between these two molecules. Additionally, a GST fusion protein consisting of the predicted SH2 domain of pp76 precipitates two tyrosine phosphoproteins from Jurkat cell lysates, and antiserum directed against phospholipase C-gamma 1 coprecipitates a tyrosine phosphoprotein with an electrophoretic mobility identical to that of pp76. These results demonstrate that this novel protein, which we term SLP-76 (SH2 domain-containing Leukocyte Protein of 76 kDa), is likely to play an important role in TCR-mediated intracellular signal transduction.
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PMID:Molecular cloning of SLP-76, a 76-kDa tyrosine phosphoprotein associated with Grb2 in T cells. 770 37

The insulin receptor, as a consequence of ligand binding, undergoes autophosphorylation of critical tyrosyl residues within the cytoplasmic portion of its beta-subunit. The 85 kDa regulatory subunit of phosphatidylinositol (PI) 3-kinase (p85), an SH2 domain protein, has been implicated as a regulatory molecule in the insulin signal transduction pathway. For the present study, glutathione S-transferase (GST) fusion proteins of p85 SH2 domains were used to determine if such motifs associate directly with the autophosphorylated human insulin receptor. The p85 N + C (amino plus carboxyl) SH2 domains were demonstrated to associate with the autophosphorylated beta-subunit, while neither the GTPase activator protein (GAP) N SH2 domain nor the phospholipase C-gamma 1 (PLC gamma 1) N + C SH2 domains exhibited measurable affinity for the activated receptor. The p85 N SH2 domain demonstrated weak association with the insulin receptor, while the p85 C SH2 domain alone formed no detectable complexes with the insulin receptor. The association of p85 N + C SH2 domains with the autophosphorylated receptor was competed efficiently by a 15-residue tyrosine-phosphorylated peptide corresponding to the carboxyl-terminal region of the insulin receptor, but not by phosphopeptides of similar length derived from the juxtamembrane or regulatory regions. The insulin receptor C domain phosphopeptide inhibited the p85 N + C SH2 domain-insulin receptor complex with an IC0.5 of 2.3 +/- 0.35 microM, whereas a 10-residue phosphopeptide derived from the insulin receptor substrate 1 (IRS-1) competed with an IC0.5 of 0.54 +/- 0.10 microM. These results demonstrate that, in vitro, there is an association between the p85 regulatory protein and the carboxyl-terminal region of the activated insulin receptor that requires the presence of both the N and C SH2 domains. Furthermore, formation of the p85/insulin receptor complex may lead to signaling pathways independent of IRS-1.
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PMID:In vitro association of the phosphatidylinositol 3-kinase regulatory subunit (p85) with the human insulin receptor. 856 77

The protein tyrosine kinase p72syk (Syk) is expressed in a variety of hematopoietic cell types, including B cells, thymocytes, mast cells and others. Both the activity and phosphotyrosine content of this enzyme increase in these cells in response to engagement of the appropriate cell surface receptors. Herein, we describe the cloning of murine Syk and its expression in Sf9 cells as a catalytically active protein. Full-length Syk and a catalytically active 42.5 kDa carboxyl terminal fragment were also expressed as glutathione S-transferase fusion proteins. Comparative reverse phase HPLC and 40% alkaline gel analysis of tryptic digests of phosphorylated Syk demonstrated that all of the major sites of autophosphorylation were also present in GST-Syk and all but one were contained in the 42.5 kDa fragment. The sites of autophosphorylation were identified using a combination of Edman sequencing and mass spectrometric analysis. Ten sites were identified. One site is located in the amino terminal half of the molecule between the two tandem Src homology 2 (SH2) domains. Five sites are located in the hinge region located between the carboxyl terminal SH2 domain and the kinase domain. Two sites lie in the kinase domain within the catalytic loop and two near the extreme carboxyl terminus. Sequences of phosphorylation sites located within the hinge region predict that Syk serves as a docking site for other SH2 domain-containing proteins. Consistent with this prediction, autophosphorylated Syk efficiently binds the carboxyl terminal SH2 domain of phospholipase C-gamma 1.
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PMID:Identification of the major sites of autophosphorylation of the murine protein-tyrosine kinase Syk. 904 38