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:3.1.4.1 (phosphodiesterase)
18,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The major aims of this study were to determine the degree of phospholipid asymmetry and the neighbor analysis of phospholipids in different types of cell membranes. For this study a penetrating probe (FDNB), a non-penetrating probe (TNBS) and a cross-linking probe (DFDNB) were used. The reaction of hemoglobin, membrane protein and membrane PE and PS of erythrocytes with DFNB and TNBS was studied over a concentration range of 0.5 to 10 mM probe. TNBS reacts to an extremely small extend with hemoglobin over the concentration range 0.4 to 4 mM whereas FDNB reacts with hemoglobin to a very large extent (50 fold more than TNBS). The reaction of membrane protein of intact erythrocytes reaches a sharp plateau at 1 mM TNBS whereas the reaction of membrane protein goes to a much larger extent with FDNB with no plateau seen up to 4 mM FDNB. This data shows that TNBS does not significantly penetrate into the cell under our conditions whereas FDNB does penetrate into the cell. The results show that there are four fold more reactive sites on proteins localized on the inner surface of the erythrocyte membrane as compared to the outer surface. TNBS at 0.5 to 2 mM concentration does not label membrane PS and labels membrane PE to a small extent. The reaction of PE with TNBS shows an initial plateau at 2 mM probe and a second slightly higher plateau between 4 to 10 mM probe. TNBS from 0.5-2.0 mM does not react with PS, but between 3 to 10 mM concentration, a very small amount of PS reacts with TNBS. Hence above 2 mM TNBS or FDNB a perturbation occurs in the membrane such that more PE and PS are exposed and react with these probes. These results demonstrate that essentially no PS is localized on the outer surface of the membrane and only 5% of the total membrane PE is localized on the outer surface of the erythrocyte membrane. TNBS and FDNB were reacted with yeast, E. coli, and Acholeplasma cells. With yeast cells, FDNB reacts to a much larger extent with PE than does TNBS, indicating that FDNB penetrates into the cell and labels more PE molecules. With E. coli, but not with erythrocytes or yeast cells, phospholipase A activity was very pronounced at pH 8.5 giving rise to a large amount of DNP-GPE from DNP-PE. A phosphodiesterase was also present which hydrolyized DNP-GPE to DNP-ethanolamine. The multilayered structure of the E. coli cell envelop did not permit a definitive interpretation of the results. It is clear, however, that TNBS and FDNB react to a different extent with PE in this cell. The Acholeplasma membrane had no detectable PE or PS but contains amino acid esters of phosphatidylglycerol. The reaction of these components with TNBS and FDNB indicate that these aminoacyl-PG are localized on both surfaces of the membrane, with 31% being on the outer surface and 69% on the inner surface...
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PMID:Differential reaction of cell membrane phospholipids and proteins with chemical probes. 0 64

alpha- and beta-Fibrinogenases (EC 3.4.21.5) were purified from Trimeresurus mucrosquamatus venom by the technique of recycling chromatography. Both enzymes were single polypeptide chains and homogeneous as judged by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis and ultracentrifugation. The sedimentation constants of alpha- and beta-fibrinogenases were 2.52 and 3.04 respectively. The molecular weight of alpha-fibrinogenase was 21 500--23 400, and that of beta-fibrinogenase was 25 000--26 000. The contents of proline, glycine and tryptophan were higher in beta-fibrinogenase than in alpha-fibrinogenase. The isoelectric points of alpha- and beta-fibrinogenases were pH 8.1 and 5.7 respectively. The optimal pH of alpha-fibrinogenase was about 7.4 and that of beta-fibrinogenase was around 8.5. The activity of alpha-fibrinogenase was completely destroyed after 30 min at 60 degrees C, pH 5.6, 7.4 and 9.0, while that of beta-fibrinogenase was not significantly affected by the same treatment. Both enzymes showed proteolytic activities toward fibrinogen and casein, but were devoid of phospholipase A, alkaline phosphomonoesterase and phosphodiesterase activities of the crude venom. The tosyl-L-arginine methylester esterase activity of beta-fibrinogenase was about 17 times that of the crude venom, while alpha-fibrinogenase was completely devoid of this activity. The fibrinogenolytic activity of alpha-fibrinogenase was markedly inhibited by EDTA and cysteine, while that of beta-fibrinogenase was inhibited markedly by phenylmethane sulfonylfluoride and slightly by tosyl-L-lysine chloromethylketone and cysteine.
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PMID:Physicochemical properties of alpha- and beta-fibrinogenases of Trimeresurus mucrosquamatus venom. 1 16

Bis(monoacylglycero)phosphate was purified from the livers of chloroquine-treated rats and labeled with tritium by a nonreductive catalytic exchange procedure. The mechanism of its degradation by rat liver lysosomes has been examined. A substantial amount of bis(monoacylglycero)P is degraded to monoglyceride and lysophosphatidic acid by a lysosomal phosphodiesterase having an acid pH optimum. Some bis(monoacylglycero)P is degraded to lysophosphatidylglycerol by lysosomal phospholipase A. In contrast, other phosphoglycerides have been reported to be degraded by sequential deacylation in lysosomes. The initial rate of breakdown of bis(monoacylglycero)P is only 10% of the rate observed for dioleoylphosphatidylcholine. [3H]Lysophosphatidylglycerol conversion to [3H]bis(monoacylglycero)P is stimulated by unlabeled bis(monoacylglycero)P, resulting in a futile cycle which allows the resynthesis of bis(monoacylglycero)P from its breakdown product, lysophosphatidylglycerol. This futile cycle and the unusual sn-1-glycerophospho-sn-1'-glycerol stereoconfiguration of the water-soluble backbone (Joutti, A., Brotherus, J., Renkonen, O., Laine, R., and Fischer, W. (1976) Biochim. Biophys. Acta 450, 206-209) may be important factors in the marked resistance of bis(monoacylglycero)P to degradation by lysosomal acid hydrolases.
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PMID:Degradation of bis(monoacylglycero)phosphate by an acid phosphodiesterase in rat liver lysosomes. 3 89

By means of DEAE-Sephadex A-50 Column chromatography, Trimeresurus gramineus venom was separated into twelve fractions. The fibrinogenolytic activities were distributed in Fractions 1 and 10. These enzymes were further purified by gel filtration and were homogeneous as judged by cellulose acetate membrane, sodium dodecyl sulfate polyacrylamide gel electrophoresis and ultracentrifugal analysis. Both of them were single peptide chains. The sedimentation constants of alpha- (Fraction 1) and beta-fibrinogenases (Fraction 10) were 2.20 and 3.60, respectively. The molecular weights of alpha- and beta-fibrinogenases were 23 500 and 25 000 respectively. The contents of proline and glycine were higher in beta-fibrinogenase than in alpha-fibrinogenase. The isoelectric points of alpha-fibrinogenase and beta-fibrinogenase were pH greater than 10 and 4.5, respectively. The optimal pH of alpha-fibrinogenase was approx. 7.4 and that of beta-fibrinogenase was approx. 9.0. The activity of alpha-fibrinogenase was completely destroyed after 30 min at 60 degrees C, pH 5.4, 7.4 and 9.0, while that of beta-fibrinogenase was much less affected by the same treatment. The specific fibrinogenolytic activity alpha-fibrinogenase was 31 mg fibrinogen/min per mg protein, while that of beta-fibrinogenase was 9 mg fibrinogen/min per mg protein. alpha-Fibrinogenase cleaved specifically the alpha(A) chain of monomeric fibrinogen without cleaving the beta(B) chain and gamma-chain. beta-fibrinogenase preferentially cleaved the beta(B) chain, and the alpha(A) chain was also partially cleaved by beta-fibrinogenase, if the incubation time was prolonged. Both enzymes showed proteolytic activities toward fibrinogen, fibrin and casein, but were devoid of phospholipase A, alkaline phosphomonoesterase and phosphodiesterase activities found in the crude venom. The tosyl-L-arginine methylester esterase activity of beta-fibrinogenase was about 14 times that of crude venom, while alpha-fibrinogenase was completely devoid of this activity. The fibrinogenolytic activity of alpha-fibrinogenase was markedly inhibited by EDTA and cysteine, while that of beta-fibrinogenase was inhibited markedly by phenylmethanesulfonylfluoride. alpha- and beta-fibrinogenases exert their fibrinogenolytic activity by a direct action on fibrinogen or fibrin without activation of plasminogen.
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PMID:Alpha and beta-fibrinogenases from Trimeresurus gramineus snake venom. 4 82

The purpose of this study was to elucidate the mechanisms by which arachidonic acid activates guanylate cyclase from guinea pig lung. Guanylate cyclase activities in both homogenate and soluble fractions of lung were examined. Guanylate cyclase activity was determined by measuring formtion of [32-P] cyclic GMP from alpha-[32-P] GTP in the presence of Mn2+, a phosphodiesterase inhibitor and a suitable GTP regenerating system. Arachidonic acid, and to a slight extent dihomo-gamma-linolenic acid, activated guanylate cyclase in homogenate but not soluble fractions. Similarly, phospholipase A2 activated homogenate but not soluble guanylate cyclase. Methyl arachidonate, linolenic, linoleic and oleic acids did not activate guanylate cyclase in either fraction. High concentrations of indomethacin, meclofenamate and aspirin inhibited activation of homogenate guanylate cyclase by arachidonic acid and phospholipase A2, without altering basal enzyme activity. These data suggested that a product of cyclooxygenase activity, present in the microsomal fraction, may have accounted for the capacity of arachidonic acid to activate homogenate guanylate cyclase. This view was supported by the findings that addition of the microsomal fraction to be soluble fraction enabled arachidonic acid to activate soluble guanylate cyclase, an effect which was reduced with cycloooxygenase inhibitors. Lipoxygenase activated guanylate cyclase in homogenate and soluble fractions. Arachidonic acid potentiated the activation of soluble guanylate cyclase by lipoxygenase, and this effect was inhibited with nordihydroguairetic acid, 1-phenyl-3-pyrazolidone and hydroquinone, but not with high concentrations of indomethacin, meclofenamate or aspirin. These data suggest that arachidonic acid activates guinea pig lung guanylate cyclase indirectly, via two independent mechanisms, one involving the microsomal fraction and the other involving lipoxygenase.
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PMID:Arachidonic acid activation of guinea pig lung guanylate cyclase by two independent mechanisms. 4 57

We have perfused isolated rat livers with hypocalcemic (4.4 mg 100 ml) Krebs-Ringer bicarbonate albumin buffer. After 15 min of perfusion, a substance appeared in the perfusate which decreased rat renal adenylate cyclase activation by parathyroid hormone (PTH). The material in the perfusate was purified greater than 50,000-fold by Bio-Gel P-10 chromatography. The purified antagonist decreased the activation of rat renal cortical adenylate cyclase by PTH, glucagon, and epinephrine 75 to 100%. Concentration response curves for each of the hormones indicated a noncompetitive interaction of the inhibitor with the hormone. The inhibition was not species-specific, as the activation of the parathyroid hormone-responsive adenylate cyclase in cat renal cortex was also abolished by the inhibitor from the perfused rat liver. The inhibitor is a peptide, Mr equal to similar to 1000, which is heat-stable, acid-stable, alkai-labile, and is destroyed by trypsin, leucine aminopeptidase, and elastase. It is not destroyed by phosphodiesterase, 5'-nucleotidase, alkaline phosphatase, neuraminidase, RNase, or phospholipase A. The inhibitor is not produced by isolated rat livers perfused with normocalcemic perfusion media. It is unclear whether the peptide is synthesized by the liver or whether it is a breakdown product of a larger peptide or protein in the liver. This is the first reported peptide inhibitor of adenylate cyclase.
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PMID:Isolation of a unique peptide inhibitor of hormone-responsive adenylate cyclase. 16 24

1. Prostaglandin synthase activity (EC 1.14.99.1) was demonstrated in bovine thyroid homogenates. 2. The synthase was characterized and shares many characteristics of the well-studied seminal vesicle enzyme and can be inhibited by indomethacin and eicosa-5,8,11,14-tetraynoic acid. 3. The enzyme is localized in the microsomal fraction and is probably associated with the plasma membranes. 4. Thyrotropin, but no other hormone tested, increased the activity of the enzyme when added to a microsomal fraction obtained from bovine thyroid. This effect is tissue-specific since thyrotropin has no effect on bovine seminal esicle or lung prostaglandin synthase. 5. Thyrotropin, cyclic AMP and the phosphodiesterase inhibitors, theophylline and quazodine increase enzyme activity when preincubated with bovine thyroid slices. 6. EDTA, when included in the pre-incubation mixture, enhances the thyrotropin effect on the enzyme but not the cyclic AMP, theophylline, or quazodine augmentation of enzyme activity in bovine thyroid slices. This suggests that phospholipase A is involved in the thyrotropin stimulation of prostaglandin formation.
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PMID:Further characterization of bovine thyroid prostaglandin synthase. 16 23

Trifluoperazine inhibits the activation of phosphodiesterase by binding to the calcium-dependent activator. To determine further the specificity by which trifluoperazine binds to activator, we compared the binding of trifluoperazine to activator prepared from several species and tissues and to a number of other calcium-binding proteins devoid of activator activity. Trifluoperazine binds to activator prepared from human, bovine, rat and rabbit brain and from chick embryo fibroblasts. In each case, the binding of trifluoperazine to activator was qualitatively similar and related quantitatively to the ability of the preparation to activate phosphodiesterase. Of the other calcium-binding proteins examined, namely, troponin-C, S-100 protein, phospholipase A, phospholipase B and myosin light chain, only troponin-C displayed any significant calcium-specific binding of trifluoperazine. The binding to troponin-C, however, appeared to be different from the binding to activator; whereas the binding of trifluoperazine to actovator showed no cooperativity, the binding to troponin-C showed positive cooperatively. These results and earlier data showing that trifluoperazine fails to bind to a variety of other proteins, indicate that the binding of trifluoperazine to the calcium-dependent activator of phosphodiesterase is selective and suggest that this binding may explain some of the biochemical and pharmacological actions of this antipsychotic agent.
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PMID:Specificity of the binding of trifluoperazine to the calcium-dependent activator of phosphodiesterase and to a series of other calcium-binding proteins. 20 50

E. coli cells were reacted with TNBS in bicarbonate-NaCl buffer, pH 8.5 (buffer A) and in phosphate-NaCl buffer, pH 7.0 (buffer B). In buffer A, DNP-GPE is the major product when FDNB is used. DNP-PE and DNP-LPE are formed in lesser amounts. Phospholipase A activity is high in buffer A. When TNBS is used, the labeling of the lipid components is less than with FDNB and more TNP-PE is formed relative to TNP-GPE. This data suggests that the phospholipases which are located primarily on the outer L-membrane of the cell wall act to a lesser extent on TNP-PE than on DNP-PE. E. coli cells were prelabeled with TNBS and FDNB in buffer A, washed and incubated in buffer A. The endogenous labeled DNP-PE gradually decreased with time with a concomitant increase in DNP-LPE and DNP-GPE due to phospholipase A activity. In contrast, the endogenous labeled TNP-PE also decreased with time as did the endogenous labeled TNP-LPE but a new orange lipid was produced. This lipid is believed to be a derivative of TNP-PE in which one of the nitro groups has been reduced to an amino group by nitroreductase. E. coli cells were prelabeled with TNBS and FDNB in buffer A, washed and incubated in buffer B. Under these conditions with both TNBS and FDNB there is an increase in TNP-PE and DNP-PE with a concomitant decrease in TNP-LPE, TNP-GPE, DNP-LPE and DNP-GPE. These results show that at neutral pH acylation occurs to regenerate TNP-PE and DNP-PE. E. coli cells were incubated with exogenous DNP-GPE or TNP-GPE in buffer A. The DNP-GPE and TNP-GPE were rapidly hydrolyzed by a phosphodiesterase to DNP-ethanolamine and TNP-ethanolamine. An orange derivative was formed which was provisionally identified as a derivative of DNP-ethanolamine or TNP-ethanolamine in which a nitro group has been reduced to an amino group by nitroreductase. The phospholipases and acylating enzymes present in the cell wall of E. coli are active on the dinitrophenyl and trinitrophenyl derivatives of PE and LPE and may act in concert to model and repair the plasma membrane.
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PMID:Interaction of membrane aminophospholipids of E. coli with fluorodinitrobenzene and trinitrobenzenesulfonate. 32 84

By means of CM-Sephadex column chromatography, Trimeresurus mucrosquamatus venom was separated into 20 fractions. Fraction XX had the marked anticoagulant action. This fraction was refractionated three times on Sephadex G-75, and a single peak was obtained. The patterns of microzone and disc electrophoresis also showed a single band. A single, symmetrical boundary with a value of 1.61 S was obtained by ultracentrifugation. It was a single peptide chain with a molecular weight of 11 700. The isoelectric point was higher than pH 10. The anticoagulant principle possessed phospholipase A activity and was calcium ion dependent. It did not possess proteolytic, tosyl-L-arginine methyl ester esterase, phosphodiesterase and alkaline phosphomonoesterase activities of the crude venom. The phospholipase A activity was heat-labile at pH 7.4, but was heat-stable at pH 5.6. The anticoagulant activity was more resistant to heat treatment as compared with phospholipase A activity. The anitoagulant action of the purified principle was competitively inhibited by platelet phospholid, tissue thromboplastin and cephalin, and was neutralized by antiserum. The anticoagulant principle inhibited platelet aggregation induced by ADP. It did not destroy fibrinogen, Factor X, prothrombin and thrombin; nor did it induce fibrinolysis nor interfere with the interaction between thrombin and fibrinogen. It is concluded that the anticoagulant action of this phospholipase A was due to the inhibition of the activations of Factors X and II through the inactivation of the procoagulant activity of phospholipids mediated partly by phospholipid-binding activity of this venom enzyme and partly by its enzymatic hydrolysis of phospholipids.
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PMID:Purification and characterization of the anticoagulant principle of Trimeresurus mucrosquamatus venom. 66 29


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