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)

Highly purified Sendai virus contained a protein kinase activity which atatlysed the phosphorylation of endogenous polypeptides or exogenous protamine sulphate. The virus contained very low levels of phosphoprotein phosphatase activity. Polyacrylamide gel analysis of the reaction product indicated that the phosphorylation was specific for certain polypeptides and varied according to whether the virus was grown in eggs or in tissue culture. This variation was partially associated with the difference in the polypeptide pattern that occurred when the virus was grown in eggs or in tissue culture. Characterization of these phosphoproteins demonstrated that the phosphate was incorporated predominantly in a phosphoester linkage with theonine residues. Using a detergent and high salt solubilization procedure, the protein kinase activity was found associated within glycoprotein free virus particles but not with the nucleocapsid-associated polypeptides. In vivo phosphorylation occurred when Sendai virus was grown in eggs or in tissue culture with [32P] and the phosphorylated polypeptides were similar to those of the protein kinase reaction product. Phosphorylation could also be detected in the infected cell and could occur once the virus particle polypeptides were being synthesized. The non-structural polypeptides were not phosphorylated.
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PMID:The phosphorylation of sendai virus proteins by a virus particle-associated protein kinase. 23 97

A systematic study of protein kinase activity and phosphorylation of membrane proteins by ATP was carried out with vesicular fragments of longitudinal tubules (light SR) and junctional terminal cisternae (JTC) derived from skeletal muscle sarcoplasmic reticulum (SR). Following incubation of JTC with ATP, a 170,000-Da glycoprotein, a 97,500-Da protein (glycogen phosphorylase), and a 55,000-60,000-Da doublet (containing calmodulin-dependent protein kinase subunit) underwent phosphorylation. Addition of calmodulin in the presence of Ca2+ (with no added protein kinase) produced a 10-fold increase of phosphorylation involving numerous JTC proteins, including the large (approximately 450,000 Da) ryanodine receptor protein. Calmodulin-dependent phosphorylation of the ryanodine receptor protein was unambiguously demonstrated by Western blot analysis. The specificity of these findings was demonstrated by much lower levels of calmodulin-dependent phosphorylation in light SR as compared to JTC, and by much lower cyclic AMP dependent kinase activity in both JTC and light SR. These observations indicate that the purified JTC contain membrane-bound calmodulin-dependent protein kinase that undergoes autophosphorylation and catalyzes phosphorylation of various membrane proteins. Protein dephosphorylation was very slow in the absence of added phosphatases, but was accelerated by the addition of phosphatase 1 and 2A (catalytic subunit) in the absence of Ca2+, and calcineurin in the presence of Ca2+. Therefore, in the muscle fiber, dephosphorylation of SR proteins relies on cytoplasmic phosphatases. No significant effect of protein phosphorylation was detected on the Ca2(+)-induced Ca2+ release exhibited by isolated JTC vesicles. However, the selective and prominent association of calmodulin-dependent protein kinase and related substrates with junctional membranes, its Ca2+ sensitivity, and its close proximity to the ryanodine and dihydropyridine receptor Ca2+ channels suggest that this phosphorylation system is involved in regulation of functions linked to these structures.
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PMID:Specific association of calmodulin-dependent protein kinase and related substrates with the junctional sarcoplasmic reticulum of skeletal muscle. 216 64

Studies of in vitro models demonstrate that a forward motility protein (FMP) is required for the initiation of forward motility in the immature epididymal spermatozoa. FMP is a heat-stable glycoprotein derived from epididymal plasma. During the epididymal maturation of spermatozoa in vivo, there is a marked increase of intrasperm pH and level of cyclic adenosine monophosphate (cAMP). Several studies suggest that exogenous FMP in concert with elevated intrasperm pH and level of cAMP initiates flagellar motility during the epididymal transit of sperm. cAMP activates sperm cytosolic cAMP-dependent protein kinases, which in turn phosphorylate multiple intrasperm phosphoproteins that may regulate flagellar motility. Exogenous calcium ion activates intact sperm motility, although it inhibits motility of demembranated cells on reactivation. Occurrence of cAMP-dependent type I and II protein kinases, a novel cAMP-independent protein kinase, and a phosphoprotein phosphatase has been demonstrated on the external surface of spermatozoa. The sperm surface has a coupled-enzyme system: ecto-cAMP-independent protein kinase and phosphoprotein phosphatase that regulate the phosphorylation and dephosphorylation of endogenous sperm ectophosphoproteins. The specific activities of these ecto-enzymes increase markedly during forward progression, suggesting that they may have a role in regulating flagellar motility.
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PMID:Biochemical parameters of initiation and regulation of sperm motility. 219 32

A 10-50-fold, biphasic increase in the rate of 32Pi labeling of eIF-4E was closely correlated with the induction of protein and glycoprotein biosynthesis when resting murine splenic B lymphocytes (B cells) were activated by bacterial lipopolysaccharide or the combination of phorbol 12-myristate 13-acetate and ionomycin. The fraction of eIF-4E which was phosphorylated only increased from 46% in resting cells to 83% in lipopolysaccharide-activated cells. This discrepancy between the increase in the fraction of phosphorylated eIF-4E and the increase in 32Pi labeling suggested that the phosphoryl group of eIF-4E turns over slowly in resting B cells compared with activated cells. The turnover rate for the eIF-4E phosphate moiety in lipopolysaccharide-activated cells was rapid (t1/2 = 2 h) in comparison to the eIF-4E polypeptide chain, which did not turn over detectably in 6 h. Neither protein kinase C nor a cyclic nucleotide-dependent protein kinase appeared to be involved in eIF-4E phosphorylation in B cells, based on the observations that the metabolic labeling of eIF-4E by 32Pi was insensitive to the protein kinase inhibitors H-7 and HA1004, and that maximal labeling occurred after protein kinase C activity was "down-regulated" to very low levels in phorbol 12-myristate 13-acetate/ionomycin-activated cells. Dephosphorylation in vivo was blocked by okadaic acid (IC50 = 200 nM). These results indicate that a rapid phosphorylation-dephosphorylation of eIF-4E is associated with high translation rates during the activation of B cells, and implicate protein phosphatase-1 (or possibly-2A) in the dephosphorylation of the initiation factor.
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PMID:Increased rate of phosphorylation-dephosphorylation of the translational initiation factor eIF-4E correlates with the induction of protein and glycoprotein biosynthesis in activated B lymphocytes. 224 37

The physico-chemical properties of phosphoprotein phosphatase (EC 1.3.1.16) from bovine spleen cell nuclei were investigated. The enzyme was shown to possess a wide substrate specificity and to catalyze dephosphorylation of phosphocasein, ATP, ADP and p-nitrophenylphosphate (pNPP). The Km values for ATP, ADP and pNPP are 0.44, 0.43 and 1.25 mM, respectively. The molecular weight of the enzyme as determined by gel filtration on Sephadex G-75 and electrophoresis in polyacrylamide gel of different concentrations is approximately 33 000. SDS-polyacrylamide gel electrophoresis revealed two protein bands with Mr 12 000 and 18 000. The enzyme molecule predominantly contains acidic amino acid residues, two free SH-groups and two disulphide bonds. Phosphoprotein phosphatase is a glycoprotein with the carbohydrate content of about 22%, and has an additional absorption maximum at 560 nm. The enzyme is competitively inhibited by ammonium molybdate (Ki = 0.37 microM) and non-competitively by sodium fluoride (Ki = 1.3 mM). Incubation of phosphoprotein phosphatase with 2 mM phenylmethylsulfonylfluoride (PMSF) for 25 hours resulted in a approximately 46% loss of the enzyme activity. Ammonium molybdate, sodium fluoride and PMSF reversibly inhibit the enzyme. Modification of aminoacid SH-groups, NH2-groups and histidine led to a decrease of the enzyme activity. Incubation of phosphoprotein phosphatase with [gamma-33P]ATP resulted in the incorporation of 0.33 mol of 33P per mol of the enzyme. The mechanism of the enzyme-catalyzed hydrolysis of the phosphoester bond is discussed.
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PMID:[Phosphoprotein phosphatases from cell nuclei of the bovine spleen: physico-chemical properties]. 299 59

An acid phosphatase species which is activated by Fe2+ was purified 3,700-fold from rat spleen by chromatography on columns containing Blue-Sepharose, concanavalin A-Sepharose, Sephadex G-100, and CM-Sephadex. The enzyme hydrolyzed aryl phosphates, nucleoside di- and triphosphates, phosphoproteins, and thiamine pyrophosphate with Km values of 10(-4) to 10(-3) M at an optimal pH of 5.0-5.8. Co-purification of the acid phosphatase and acid phosphoprotein phosphatase indicated that they were identical. The purified enzyme was glycoprotein in nature, showing four heterogeneous forms on acid polyacrylamide gel electrophoresis (pI values, 7.8, 8.0, 8.3, and 8.5), but it gave a molecular weight of 33,000 on sodium dodecyl sulfate-gel electrophoresis and gel permeation chromatography. The enzyme had a purple color (lambda max 545 nm) and contained 2 iron atoms per enzyme molecule. Among reductants, ascorbic acid and Fe2+ were the best activators, although their combined effect was not additive. Fe2+ and ascorbic acid both changed the purple enzyme into the same active form (lambda max 515 nm), giving almost the same kinetic constants for substrates and for inhibitors such as molybdate, phosphate and fluoride. However, low concentrations of Fe2+, from 0.01 mM to 1.0 mM, immediately and reversibly activated the enzyme, whereas high concentrations of ascorbic acid over 1 mM were required for maximal activation, which was slow and irreversible.
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PMID:Purification and characterization of a purple acid phosphatase from rat spleen. 670 19

Alkaline phosphatase (E.C.3.1.3.1.) from unerupted bovine pulp was extracted from the microsomal fraction with eta-butanol and purified 77-fold, using DEAE-cellulose chromatography, Sephadex G-200 gel-filtration and concanavalin-A affinity chromatography, to a final specific activity of 92.3 units/mg protein. Affinity chromatography confirmed the glycoprotein nature of the enzyme. The pH optimum for the purified enzyme was 10.0 with rho-nitrophenylphosphate, and 8.7 with phosphoserine. The apparent Km was estimated to be 0.7 mM, using rho-nitrophenylphosphate in glycine-NaOH buffer, pH 10.0. The enzyme was markedly inhibited by EDTA, bromotetramisole and homoarginine but was insensitive to phenylalanine, and therefore resembled the alkaline phosphatase of liver and bone, but not that of intestine and placenta. No protein phosphatase activity towards dentine phosphoprotein and phosvitin was observed.
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PMID:Purification and properties of bovine dental-pulp alkaline-phosphatase. 695 31

Tissue non-specific alkaline phosphatase is a membrane-bound glycoprotein enzyme which is characterized by its phosphohydrolytic, protein phosphatase, and phosphotransferase activities. This enzyme is distributed virtually in all mammalian tissues, particularly during embryonic development. Its expression is stage-specific and can be demonstrated in the developing embryo as early as the 2-cell stage. It has been suggested that tissue non-specific alkaline phosphatase might play a role in tissue formation. In the study reported here, a gene-transfer approach was employed to investigate possible roles for this enzyme by inserting the cDNA for rat tissue non-specific alkaline phosphatase into CHO and LLC-PK1 cells. Permanently transfected cell-lines expressing varying levels of alkaline phosphatase were established. The data showed that functional enzyme was expressed in the transfected cells. Cell spreading and attachment were enhanced in transfected CHO cells expressing high levels of tissue non-specific alkaline phosphatase but not in the LLC-PK1 cells. Further, in CHO cells, proliferation was shown to be inversely proportional to the level of the tissue non-specific alkaline phosphatase expression. Homotypic cell association was demonstrated in both alkaline phosphatase-positive and alkaline phosphatase-negative cells in both CHO and LLC-PK1 cell-lines. Taken together, these findings suggest that in addition to a role in mineralization of bone, tissue non-specific alkaline phosphatase might also play a role in other cell activities, including those related to differentiation, such as cell-cell or cell-substrate interaction and proliferation.
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PMID:Changes in cell adhesion and cell proliferation are associated with expression of tissue non-specific alkaline phosphatase. 750 6

When platelets are stimulated by the addition of thrombin, a series of temporally linked signaling events are initiated. Some of the early events are needed to engage the integrin glycoprotein (GP) IIb-IIIa in a high-affinity state. This in turn leads to aggregation, which initiates a wave of events distinct from those triggered by thrombin. Platelet responses are sensitive to protein serine/threonine phosphatase inhibitors, but which events are dependent on protein phosphatase activity is not known. In the present studies, the effect of the phosphatase inhibitor calyculin A on aggregation-induced signaling was examined. The addition of 0.2 unit/mL thrombin caused aggregation-dependent redistribution of cytoskeletal proteins (actin binding protein, talin, vinculin, and alpha-actinin), glycoproteins (GPIIb-IIIa, PECAM), and signaling molecules (PI3-kinase, pp60c-src) to the cytoskeletal fraction of platelets. Addition of 1-2 microM calyculin A blocked the ability of 0.2 unit/mL thrombin to induce aggregation and the association of these molecules with the cytoskeleton. Aggregation (60-80% of control) was restored if 1 unit/mL thrombin was added, but there was no corresponding redistribution of actin binding protein, talin, vinculin, alpha-actinin, GPIIb-IIIa, PECAM, PI3-kinase, and pp60c-src to the cytoskeleton. Treatment of platelets with calyculin A resulted in an increase in the phosphorylation state of a membrane skeletal protein of 50 kDa. These data strongly suggest that platelet aggregation is dissociable from aggregation-induced signaling, which is dependent on type 1 and 2A phosphatase activities.
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PMID:Aggregation-dependent signaling in human platelets is sensitive to protein serine/threonine phosphatase inhibitors. 762 26

Mouse malignant T-lymphoma CS-21 cells grow in vitro in the presence of CA-12 stromal cells, but they undergo apoptotic cell death with DNA fragmentation when cultured alone. Because apoptosis of CS-21 cells was not inhibited by soluble factors secreted from CA-12 stromal cells, cell-cell interactions between the two seemed to be important to inhibit apoptosis. We found that CS-21 cell adhesion was mediated by M(r) 168,000 and M(r) 23,000 proteins and that apoptosis-inhibitory signals were transmitted through these proteins. In this study, we identified the M(r) 23,000 cell adhesion molecule as a glycosylphosphatidylinositol-anchored Thy-1 (CD90) glycoprotein. Cross-linking of M(r) 23,000 protein with anti-M(r) 23,000 mAb and a second antibody transiently raised the [Ca2+]i and activated calcineurin in CS-21 cells, as has been observed in normal T lymphocytes stimulated by cross-linking anti-Thy-1 mAbs. However, differing from normal T lymphocytes, CS-21 cells could grow either by the transient increase in [Ca2+]i or by the activation of protein kinase C. Furthermore, M(r) 23,000 protein-mediated cell survival of CS-21 cells was not accompanied by expression of the apoptosis-inhibiting protein bcl-2, although protein kinase C-activated cell survival was attended by bcl-2 expression. These results indicate that the M(r) 23,000 protein (Thy-1) of CS-21 lymphoma cells functions as a cell adhesion molecule capable of transducing signals of cell survival and growth that are not followed by bcl-2 expression.
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PMID:Apoptosis inhibition by anti-M(r) 23,000 (Thy-1) monoclonal antibodies without inducing bcl-2 expression. 779 3


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