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:3.1.3.16 (calcineurin)
17,112 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two Tyr-protein phosphatase inhibitors, termed inhibitor H (Mr greater than 500,000) and inhibitor L (Mr 38,000), have been detected in bovine brain extracts. The inhibitors were partially purified by chromatography on DEAE-cellulose and Sephacryl S-300. Both inhibitors are proteins, as judged by their inactivation by proteinase K, and they exhibited remarkable stability during incubation at 95 degrees C. Of seven Tyr-protein phosphatase activities that we have isolated from bovine brain, PTP-4 and PTP-5 were most sensitive to the inhibitor proteins. Inhibition of the other five Tyr-protein phosphatases was only observed at very high inhibitor concentrations. The IC50 values for the inhibition of PTP-4 by inhibitor H and inhibitor L were 2- and 10-fold higher than those for the inhibition of PTP-5. Inhibition of PTP-5 by either inhibitor was rapid (maximum effect in less than 1 min) and readily reversed upon removal of the inhibitors by dilution. Inhibitor H and inhibitor L are distinct from the three heat-stable protein inhibitors of Ser/Thr-protein phosphatase 1. The ability of inhibitor H and inhibitor L to preferentially inhibit PTP-4 and PTP-5 provides an important new criterion that can be used to distinguish these enzymes from other Tyr-protein phosphatases. The two inhibitor proteins may be involved in regulating the activity of PTP-4 and PTP-5.
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PMID:Phosphotyrosyl-protein phosphatases. II. Identification and characterization of two heat-stable protein inhibitors. 246 74

Activation of glycogen synthase is one of the major metabolic events triggered by exposure of cells to insulin. The molecular mechanism by which insulin activates glycogen synthase was investigated. The possible role of Ras and mitogen-activated protein kinase cascade was investigated with a stable cell line, CHO-IR-C/S 46, that overexpresses insulin receptors and a catalytically inactive SH-PTP 2 protein phosphatase and in which insulin does not induce the formation of the Ras-GTP complex or the subsequently activation of the mitogen-activated protein kinase cascade. Insulin activated glycogen synthase in this cell line to a similar extent as in parental CHO-IR cells. The importance of heteromeric phosphoinositide (PI) 3-kinase in insulin activation of glycogen synthase was examined in a stable cell line, CHO-IR/delta p85, that overexpresses insulin receptors and a dominant negative mutant (delta p85) of the 85-kDa subunit of PI 3-kinase that lacks the binding site for the catalytic 110-kDa subunit. Insulin-dependent activation of PI-3 kinase and glucose transport, but not the formation of the Ras-GTP complex, are markedly attenuated in this cell line. In CHO-IR/delta p85 cells, insulin activated glycogen synthase to a similar extent as in parental CHO-IR cells. The failure of overproduction of the mutant (delta p85) protein to inhibit insulin activation of glycogen synthase was also confirmed by transient expression in Rat 1 cells with the use of a recombinant vaccinia virus. However, wortmannin abolished insulin activation of glycogen synthase in all cell lines. These data suggest that existence of a Ras-independent and wortmannin-sensitive pathway for activation of glycogen synthase by insulin.
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PMID:Ras-independent and wortmannin-sensitive activation of glycogen synthase by insulin in Chinese hamster ovary cells. 774 67

The low molecular weight phosphotyrosine-protein phosphatase (LMW-PTP) is a cytosolic phosphotyrosine-protein phosphatase specifically interacting with the activated platelet-derived growth factor (PDGF) receptor through its active site. Overexpression of the LMW-PTP results in modulation of PDGF-dependent mitogenesis. In this study we investigated the effects of this tyrosine phosphatase on the signaling pathways relevant for PDGF-dependent DNA synthesis. NIH 3T3 cells were stably transfected with active or dominant negative LMW-PTP. The effects of LMW-PTP were essentially restricted to the G1 phase of the cell cycle. Upon stimulation with PDGF, cells transfected with the dominant negative LMW-PTP showed an increased activation of Src, whereas the active LMW-PTP induced a reduced activation of this proto-oncogene. We observe that c-Src binding to PDGF receptor upon stimulation is prevented by overexpression of LMW-PTP. These effects were associated with parallel changes in myc expression. Moreover, wild-type and dominant negative LMW-PTP differentially regulated STAT1 and STAT3 activation and tyrosine phosphorylation, whereas they did not modify extracellular signal-regulated kinase activity. However, these modifications were associated with changes in fos expression despite the lack of any effect on extracellular signal-regulated kinase activation. Other independent pathways involved in PDGF-induced mitogenesis, such as phosphatidylinositol 3-kinase and phospholipase C-gamma1, were not affected by LMW-PTP. These data indicate that this phosphatase selectively interferes with the Src and the STATs pathways in PDGF downstream signaling. The resulting changes in myc and fos proto-oncogene expression are likely to mediate the modifications observed in the G1 phase of the cell cycle.
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PMID:The Src and signal transducers and activators of transcription pathways as specific targets for low molecular weight phosphotyrosine-protein phosphatase in platelet-derived growth factor signaling. 950 79

Bioactive compound(s) extracted from cinnamon potentiate insulin activity, as measured by glucose oxidation in the rat epididymal fat cell assay. Wortmannin, a potent PI 3'-kinase inhibitor, decreases the biological response to insulin and bioactive compound(s) from cinnamon similarly, indicating that cinnamon is affecting an element(s) upstream of PI 3'-kinase. Enzyme studies done in vitro show that the bioactive compound(s) can stimulate autophosphorylation of a truncated form of the insulin receptor and can inhibit PTP-1, a rat homolog of a tyrosine phosphatase (PTP-1B) that inactivates the insulin receptor. No inhibition was found with alkaline phosphate or calcineurin suggesting that the active material is not a general phosphatase inhibitor. It is suggested, then, that a cinnamon compound(s), like insulin, affects protein phosphorylation-dephosphorylation reactions in the intact adipocyte. Bioactive cinnamon compounds may find further use in studies of insulin resistance in adult-onset diabetes.
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PMID:Regulation of PTP-1 and insulin receptor kinase by fractions from cinnamon: implications for cinnamon regulation of insulin signalling. 976 7

Low molecular weight phosphotyrosine-protein phosphatase (LMW-PTP) shares no general sequence homology with other PTPs, although it has an active site sequence motif CXXXXXR and a reaction mechanism identical to those of all PTPs. The main function of this enzyme is the down-regulation of platelet-derived growth factor and insulin receptors. Both human LMW-PTP isoenzymes are inactivated by H2O2. The enzymes are protected from inactivation by Pi, a competitive inhibitor, suggesting that the H2O2 reaction is directed to active site. Analysis of free thiols performed on the inactivated enzymes demonstrates that only two out of the eight LMW-PTP cysteines are modified. Time-course high performance liquid chromatography-electrospray mass spectrometry, together with specific radiolabeling and tryptic fingerprint analyses, enables us to demonstrate that H2O2 causes the oxidation of Cys-12 and Cys-17 to form a disulfide bond. Because both residues are localized into the active site region, this modification inactivates the enzyme. Fluorescence spectroscopy experiments suggest that the fold of the enzyme is modified during oxidation by H2O2. Because a physiological concentration of H2O2 produces enzyme inactivation and considering that the activity is restored by reduction with low molecular weight thiols, we suggest that oxidative stress conditions and other processes producing hydrogen peroxide regulate the LMW-PTP in the cell.
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PMID:The inactivation mechanism of low molecular weight phosphotyrosine-protein phosphatase by H2O2. 982 91

Inspection of the genomes for the bacteria Bacillus subtilis 168, Borrelia burgdorferi B31, Escherichia coli K-12, Haemophilus influenzae KW20, Helicobacter pylori 26695, Mycoplasma genitalium G-37, and Synechocystis sp PCC 6803 and for the archaeons Archaeoglobus fulgidus VC-16 DSM4304, Methanobacterium thermoautotrophicum delta H, and Methanococcus jannaschii DSM2661 revealed that each contains at least one ORF whose predicted product displays sequence features characteristic of eukaryote-like protein-serine/threonine/tyrosine kinases and protein-serine/threonine/tyrosine phosphatases. Orthologs for all four major protein phosphatase families (PPP, PPM, conventional PTP, and low molecular weight PTP) were present in the bacteria surveyed, but not all strains contained all types. The three archaeons surveyed lacked recognizable homologs of the PPM family of eukaryotic protein-serine/threonine phosphatases; and only two prokaryotes were found to contain ORFs for potential phosphatases from all four major families. Intriguingly, our searches revealed a potential ancestral link between the catalytic subunits of microbial arsenate reductases and the protein-tyrosine phosphatases; they share similar ligands (arsenate versus phosphate) and features of their catalytic mechanism (formation of arseno-versus phospho-cysteinyl intermediates). It appears that all prokaryotic organisms, at one time, contained the genetic information necessary to construct protein phosphorylation-dephosphorylation networks that target serine, threonine, and/or tyrosine residues on proteins. However, the potential for functional redundancy among the four protein phosphatase families has led many prokaryotic organisms to discard one, two, or three of the four.
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PMID:The serine, threonine, and/or tyrosine-specific protein kinases and protein phosphatases of prokaryotic organisms: a family portrait. 986 22

PTP-S2 is a tyrosine specific protein phosphatase that binds to DNA and is localized to the nucleus in association with chromatin. It plays a role in the regulation of cell proliferation. Here we show that the subcellular distribution of this protein changes during cell division. While PTP-S2 was localized exclusively to the nucleus in interphase cells, during metaphase and anaphase it was distributed throughout the cytoplasm and excluded from condensed chromosomes. At telophase PTP-S2 began to associate with chromosomes and at cytokinesis it was associated with chromatin in the newly formed nucleus. It was hyperphosphorylated and showed retarded mobility in cells arrested in metaphase. In vitro experiments showed that it was phosphorylated by CK2 resulting in mobility shift. Using a deletion mutant we found that CK2 phosphorylated PTP-S2 in the C-terminal non-catalytic domain. A heparin sensitive kinase from mitotic cell extracts phosphorylated PTP-S2 resulting in mobility shift. These results are consistent with the suggestion that during metaphase PTP-S2 is phosphorylated (possibly by CK2 or a CK2-like enzyme), resulting in its dissociation from chromatin.
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PMID:PTP-S2, a nuclear tyrosine phosphatase, is phosphorylated and excluded from condensed chromosomes during mitosis. 1082 96

Protein tyrosine phosphatase 1B (PTP1B) attenuates insulin signaling by catalyzing dephosphorylation of insulin receptors (IR) and is an attractive target of potential new drugs for treating the insulin resistance that is central to type II diabetes. Several analogues of cholecystokinin(26)(-)(33) (CCK-8) were found to be surprisingly potent inhibitors of PTP1B, and a common N-terminal tripeptide, N-acetyl-Asp-Tyr(SO(3)H)-Nle-, was shown to be necessary and sufficient for inhibition. This tripeptide was modified to reduce size and peptide character, and to replace the metabolically unstable sulfotyrosyl group. This led to the discovery of a novel phosphotyrosine bioisostere, 2-carboxymethoxybenzoic acid, and to analogues that were >100-fold more potent than the CCK-8 analogues and >10-fold selective for PTP1B over two other PTP enzymes (LAR and SHP-2), a dual specificity phosphatase (cdc25b), and a serine/threonine phosphatase (calcineurin). These inhibitors disrupted the binding of PTP1B to activated IR in vitro and prevented the loss of tyrosine kinase (IRTK) activity that accompanied PTP1B-catalyzed dephosphorylation of IR. Introduction of these poorly cell permeant inhibitors into insulin-treated cells by microinjection (oocytes) or by esterification to more lipophilic proinhibitors (3T3-L1 adipocytes and L6 myocytes) resulted in increased potency, but not efficacy, of insulin. In some instances, PTP1B inhibitors were insulin-mimetic, suggesting that in unstimulated cells PTP1B may suppress basal IRTK activity. X-ray crystallography of PTP1B-inhibitor complexes revealed that binding of an inhibitor incorporating phenyl-O-malonic acid as a phosphotyrosine bioisostere occurred with the mobile WPD loop in the open conformation, while a closely related inhibitor with a 2-carboxymethoxybenzoic acid bioisostere bound with the WPD loop closed, perhaps accounting for its superior potency. These CCK-derived peptidomimetic inhibitors of PTP1B represent a novel template for further development of potent, selective inhibitors, and their cell activity further justifies the selection of PTP1B as a therapeutic target.
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PMID:Small molecule peptidomimetics containing a novel phosphotyrosine bioisostere inhibit protein tyrosine phosphatase 1B and augment insulin action. 1134 29

PTP-MEG2 is an intracellular protein tyrosine phosphatase with a putative lipid-binding domain at the N-terminus. The present study reports expression, purification, and characterization of the full-length form of the enzyme plus a truncated form containing the catalytic domain alone. Full-length PTP-MEG2 was expressed with an adenovirus system and purified from cytosolic extracts of human 293 cells infected with the recombinant adenovirus. The purification scheme included chromatographic separation of cytosolic extracts on fast flow Q-Sepharose, heparin-agarose, l-histidyldiazobenzylphosphonic acid agarose, and hydroxylapatite. The enrichment of PTP-MEG2 from the cytosol was about 120-fold. The truncated form of PTP-MEG2 was expressed in E. coli cells as a non-fusion protein and purified by using a chromatographic procedure similar to that used for the full-length enzyme. The purified full-length and truncated enzymes showed single polypeptide bands on SDS-polyacrylamide gel electrophoresis under reducing conditions and behaved as monomers on gel exclusion chromatography. With para-nitrophenylphosphate and phosphotyrosine as substrates, both forms of the enzyme exhibited classical Michaelis-Menten kinetics. Their responses to pH, ionic strength, metal ions, and protein phosphatase inhibitors are similar to those observed with other characterized tyrosine phosphatases. Compared with full-length PTP-MEG2, the truncated DeltaPTP-MEG2 displayed significantly higher V(max) and lower K(m) values, suggesting that the N-terminal putative lipid-binding domain may have an inhibitory role. The full-length and truncated forms of PTP-MEG2 were also expressed as GST fusion proteins in E. coli cells and purified to near homogeneity through affinity columns. However, the specific phosphatase activities of the GST fusion proteins were 10-25-fold below those obtained with the correspondent non-fusion proteins.
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PMID:Purification and characterization of protein tyrosine phosphatase PTP-MEG2. 1211 18

It has been demonstrated that signal transducer and activator of transcription-3 (STAT3) is activated after cerebral ischemia/reperfusion (I/R) in cortex and striatum. In this study, we investigated whether STAT3 was rapidly activated in hippocampus by cerebral ischemia without reperfusion in four-vessel occlusion (4-VO) model of Sprague-Dawley (SD) rats. The results showed that tyrosine phosphorylation and DNA binding activity of STAT3 was rapidly increased by ischemia. The p-STAT3 level in cytoplasm increased 5 min after occlusion and reached a peak at 10 min following ischemia (1.7 folds vs sham) by means of immunoblotting (IB). P-STAT3 in nucleus was gradually enhanced with its peak activity occurring at 30 min of ischemia (2.3 folds vs sham). Electrophoretic mobility shift assay (EMSA) with STAT3 probe demonstrated that DNA binding activity of STAT3 in nuclear extracts increased from 5 min and peaked at 30 min of ischemia (3.2 folds vs sham). These changes were prevented by genistein (a protein tyrosine kinase inhibitor) and antioxidant N-acetyl-L-cysteine (NAC), but promoted by sodium orthovanadate (a protein phosphatase inhibitor), which were administered to the SD rats 20 min before ischemia. These results indicate that the activation of STAT3 following cerebral ischemia may be modulated by PTK/PTP, and that this pathway may be of benefit to the adaptation of the hippocampal neurons to oxidative stress.
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PMID:Activation of STAT3 induced by cerebral ischemia in rat hippocampus and its possible mechanisms. 1281 99


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