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:4.6.1.1 (adenylate cyclase)
19,190 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An activating factor of adenylate cyclase (EC 4.6.1.1) HAS BEEN OBTAINED FROM DETERGENT-DISPERSED PREPARATIONS OF PORCINE CEREBRAL CORTEX BY COLUMN CHROMATOGRAPHY ON ECTEOLA-cellulose. The factor was identified by acrylamide gel electrophoresis and by enzyme activation studies as the Ca2+-binding protein that regulates the activity of a brain cyclic nucleotide phosphodiesterase. This Ca2+-binding protein confers a Ca2+-dependent activation upon the adenylate cyclase, which is reversed by the subsequent addition of egta in excess of the free Ca2+. It is proposed that this Ca2+-dependent regulator controls enzymatic activities responsible for the synthesis of adenosine 3':5'-monophosphate and for the hydrolysis of guanosine 3':5'-monophosphate.
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PMID:Identification of a calcium-binding protein as a calcium-dependent regulator of brain adenylate cyclase. 16 29

Simple one step assay methods for adenylate cyclase (ATP pyrophosphate-lyase (cyclizing) EC 4.6.1.1) and cyclic nucleotide phosphodiesterases (3',5'-cyclic nucleotide 5'-nucleotidohydrolase EC 3.1.4.17) have been developed. [alpha-32-P] ATP is used as the substrate for adenylate cyclase. Acid-heat destruction of [32-P] ATP remaining after the cyclase reaction followed by Zn-Ba treatment quantitatively leaves cyclic [32-P] AMP in the supernatant essentially free from other 32-P-containing compounds. This assay method requires no corrections for recovery and routinely yields blank values less than 0.03 per cent. If higher sensitivity is desired, a simple 5 min alumina column step can be introduced into the procedure which quantitatively elutes cyclic [32-P] AMP directly into a liquid scintillation vial and lowers the blank values to less than 0.002 per cent. This method is rapid and easily performed, without sacrificing high reliability, specificity, or sensitivity. One step phosphodiesterase assays are easily accomplished using 32-P-labeled cyclic nucleotides as substrates. Descending paper chromatography of the reaction mixture on individual 2 cm wide paper strips gives a complete and quantitative separation of all possible products including [5'-32-P] AMP and [5'-32-P] GMP from their respective 32-P-labeled 3',5'-cyclic nucleotides in 1-2 h. The paper strips are cut, inserted in scintillation vials without scintillant and the 32-P-products determined by Cerenkov counting. Low blank values of less than 0.5 per cent and the use of high specific activity 32-P-labeled cyclic nucleotide substrates make this method the most reliable and most sensitive phosphodiesterase assay described to date. Because of the simplicity, specificity, and high sensitivity obtainable with these assay methods using 32-P-labeled substrates, we have also devised simple conditions for the preparation and purification of [alpha-32-P] ATP, cyclic [32-P] AMP and cyclic [32-P] GMP with specific activities in excess of 100 Ci/mmol. These high specific activity 32-Plabeled cyclic nucleotides are important for these new assay methods and are also useful to follow purification recovery of endogenous cyclic AMP and cyclic GMP from biological materials before protein binding or radioimmunological isotope displacement assays when performed in the femtomole range.
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PMID:Assay for adenylate cyclase and cyclic nucleotide phosphodiesterases and the preparation of high specific activity 32-P-labeled substrates. 16 81

The initial rate of net glycerol release in norepinephrine-stimulated adipose tissue fragments was inhibited (40-78%) by procaine-HCl (1-5mM), whereas basal (unstimulated) lipolysis was unaffected. A dose-related inhibition of norepinephrine-induced lipolysis by procaine-HCl (0.1-1 mM) also occurred in adipocytes. Procaine-induced antilipolysis was associated with an augmented rather than a reduced hormone-stimulated increment in intracellular cyclic AMP. The dissociation of lipolysis from cyclic AMP accumulation has been termed the uncoupling effect of procaine. This effect of procaine was employed to define the precise mechanism of action of the antilipolytic drug clofibrate (Atromid-S) which inhibits lipolysis by reducing cyclic AMP. A reduction in cyclic AMP by clofibrate was demonstrated in norepinephrine-stimulated cells exposed to procaine (uncoupled system). Thus, the inhibitory effect of clofibrate on cyclic AMP could not be attributed to accumulation of products of lipolysis. Because neither procaine-HCl nor clofibrate had any effect on the low Km 3':5'-cyclic-AMP phosphodiesterase (EC 3.1.4.17) activity in hormone stimulated cells, the clofibrate-induced reduction in cyclic AMP was attributed to its direct action on adipocyte adenylate cyclase.
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PMID:Uncoupling of lipolysis from cyclic AMP by procaine: a tool for studying the mechanism of action of antilipolytic agents. 16 76

Catecholamine-sensitive adenylate cyclase, cyclic nucleotide phosphodiesterase, adenosine 3', 5'-monophosphate (cyclic AMP)-dependent protein kinase, kinase substrate, and phosphoprotein phosphatase have variously been reported to be present in preparations of myocardial cellular membranes that function in the movement of Ca2+ in and out of the cell and in intracellular Ca2+ translocations, indicating that these membranees possess the equipment for the formation and destruction of cyclic Amp as well as for the initiation, effectuation, and termination of a possible membrane action of the nucleotide. It has also been observed that phosphorylation of seryl residues of protein in sarcolemma- and sarcotubule-rich myocardial subcellular fractions by cyclic AMP activated intrinsic and extrinsic protein kinases confers upon these membran structures an enhanced ability to bind or take up Ca2+ and that dibutyryl cyclic AMP, like adrenaline, produces in intact cardiac muscle simultaneous increases in contractile force and in the uptake of extracellular Ca2+. These findings are suggestive of a second messenger role of cyclic AMP in the beta-adrenoreceptor-mediated actions of catecholamines on myocardial contractile force and relaxation, in which Ca2+ would serve as a third messenger and be subject, respectively, to more effective removal from its binding sites on troponin. An alternative interpretation regards Ca2+ and cyclic AMP as interdependent twin second messengers in the catecholamine-induced inotropism. Since the physiological meaning of the reported effects of cyclic AMP on isolated myocardial membrane preparations is far from established an instances of a dissociation between the effects of catecholamines on myocardial contractile force and cyclic AMP levels have been observed, there is still room for hypotheses that relegate cyclic AMP to a nonobligatory, at most, supportive role in the action of the catecholamines on cardiac contraction.
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PMID:Adenosine 3',5'-monophosphate, the myocardial cell membrane, and calcium. 17 10

Adenyl cyclase and cyclic nucleotide phosphodiesterase activities were assayed in homogenates of hind leg skeletal muscle from dystrophic and normal mice. Adenyl cyclase activity was stimulated 2.5 times by epinephrine and 6 times by fluoride over the basal activity in both dystrophic and normal mice. The activity of adenyl cyclase from dystrophic muscle of mice was significantly higher than that of normal mice under all the conditions tested (i.e. basal, epinephrine and fluoride). Cyclic nucleotide phosphodiesterase from skeletal muscle of mice has two Km's (2.1 and 11 mumol/l) which suggests the existence of either two forms of enzyme or a single enzyme with negative cooperativity. The activity of this enzyme was significantly elevated in the skeletal muscle of dystrophic mice compared to the normal controls. The available evidence suggests that the same cyclic nucleotide phosphodiesterase is responsible for the hydrolysis of both cyclic AMP and cyclic GMP.
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PMID:Adenyl cyclase and cyclic nucleotide phosphodiesterase activities in murine muscular dystrophy. 17 29

Changes in the activity of adenylate cyclase [EC 4.6.1.1] of serum-stimulated hamster BHK cells in culture were investigated in relation to changes in the intracellular level of adenosine 3',5'-cyclic monophosphate (cyclic AMP). Addition of calf serum to quiescent cultures decreases the activity of adenylate cyclase, followed by cellular DNA synthesis. Cyclic AMP levels drop in parallel with the decrease in adenylate cyclase activity. Kinetic analysis of cyclic AMP phosphodiesterase [EC 3.1.4.17] revealed that BHK cells contain two forms of the enzyme, one with a Km of 0.77 muM and the other with Km of 17.0 muM. The activity of these enzymes is not affected by the addition of serum to tissue culture cells. The findings indicate that the decrease in adenylate cyclase activity is directly responsible for the decrease in cyclic AMP levels that is observed immediately after serum addition. Low levels of cyclic AMP continue for several hours during serum treatment, followed by a transient increase in cyclic AMP in the early stages of cellular DNA synthesis, at which time the activity of cyclic AMP phosphodiesterase with a low Km shows a slight decrease.
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PMID:Correlation between adenosine 3',5'-cyclic monosphosphate levels, adenylate cyclase activity, and adenosine 3',5'-cyclic monophosphate phosphodiesterase activity in tissue culture cells stimulated by serum. 17 46

Exposure of platelets to 1 C led to a transient increase in cyclic AMP levels (determined either by a protein binding method or by radioimmunoassay) within five to ten minutes reaching a maximum 10 to 15 minutes after chilling was begun and returning subsequently to baseline values. Addition of EDTA to the platelet suspension medium prevented this increase. Rewarming at 37 C produced a sudden reduction in platelet cyclic AMP. To determine whether the cold-induced increase in cyclic AMP was due to a transient stimulation of platelet adenylate cyclase or a rapid inhibition of phosphodiesterase, these enzymes were assayed in ruptured platelet suspensions. Platelet adenylate cyclase activity was found to possess certain characteristics similar to those of the enzyme derived from other sources but there was a marked potentiation of fluoride-stimulated adenylate cyclase activity by 0.001 M EDTA. This effect was limited to low EDTA concentrations. Exposure of platelets to 1 C for up to 60 minutes did not increase adenylate cyclase activity but lowered it substantially compared with controls kept at room temperature. Phosphodiesterase activity at 1 C was depressed sooner and to a greater extent than was adenylate cyclase. The transient rise in cyclic AMP levels in chilled platelets appears to be due to a disproportionate reduction of cyclic nucleotide phosphodiesterase activity.
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PMID:Effect of chilling on platelet cyclic adenosine 3:5-monophosphate and adenylate cyclase activity. 17 53

The ultraviolet spectrum of a protein activator of cyclic nucleotide phosphodiesterase and adenylate cyclase purified to homogeneity from bovine brain displayed absorption peaks at 252, 259, 265, 269, and 277 nm. The activator contained no phosphate and did not serve as a substrate for cyclic adenosine 3':5'-monophosphate- or cyclic guanosine 3':5'-monophosphate-dependent protein kinases. The activator binds Ca2+, and the active form appears to be a Ca2+ activator complex (Lin, Y.M., Liu, Y.P., and Cheung, W.Y. (1974) J. Biol. Chem. 249, 4943-4954). Optical rotatory dispersion measurement showed that the Ca2+-free activator exhibited a reduced mean residue rotation ([m']231) of -5700, corresponding to 39% of helical content. In the presence of Ca2+, the [m']231 was increased to -7500, corresponding to 57% of helical content. The Ca2+ -induced conformational change was corroborated by a chemical method. In the presence of Ca2+, the activator was more resistant to trypsin inactivation, presumably because proteins with more helical structures are more resistant to tryptic attack. The activator is rich in aspartate and glutamate. Chemical block of some of the carboxyl groups with glycine ethyl ester or methoxyamine diminished the [m']231 of the activator and its activity, suggesting that blockade of some of the carboxyl groups in the activator unfolded the molecule, leading to a loss of activity. We conclude that Ca2+, which confers more helical structure to the activator, converts the inactive, less helical structure to the active, more helical structure, and that chemical modification of the activator leading to unfolding of the molecule abolishes its biological activity.
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PMID:Cyclic 3':5'-nucleotide phosphodiesterase. Ca2+ confers more helical conformation to the protein activator. 18 19

The effect of five phosphodiesterase (PDE) inhibitors (papaverine, IBMX, theophyllamine, dipyridamol and M & B 22,948) was studied on adenylate cyclase and on cyclic nucleotide phosphodiesterase activities in extracts of rat caudate nucleus. For comparison the effect on DA turnover and on turning behaviour in rats with unilateral lesions of the nigro-neostriatal DA nerurons was studied. Cyclic AMP PDE was inhibited by papaverine, dipyridamol, IBMX, M & B 22,948 and theophyllamine in that order of potency. Cylcic GMP PDE was inhibited by IBMX, papaverine, M & B 22,948 and theophyllamine, but not by dipyridamol. Basal adenylate cyclase washigher if assayed in the presence of papaverine or dipyridamol than if theophyllamine or IBMX was present. The degree of stimulation caused by DA was not significantly influenced by the PDE inhibitors. Papaverine and dipyridamol enhanced DA disappearance in the caudate nucleus and the tuberculum accumbens, but not in the median eminence. Caffeine had no significant effect. Papaverine (1-28 mg/kg) had no signigicant effect on L-dopa (5 mg/kg)-induced turning, and actually inhibited turning induced by the combination of L-dopa (10 mg/kg) and atropine (5 mg/kg). The other four PDE inhibitors all potentiated L-dopa-induced turning. Theophyllamine (20 mg/kg) and IBMX (5 mg/kg) even caused turning when given alone. The data are compatible with the opinion that PDE inhibition leads to an enhanced effect of DA in the caudate nucleus. However, the results also demonstrate that several of the PDE inhibitors have effects on central DA mechanisms that are difficult to explain solely on the basis of PED inhibition.
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PMID:Effect of some phosphodiesterase inhibitors on central dopamine mechanisms. 18 7

Depending on growth conditions, the adenosine 3':5'-cyclic monophosphate (cyclic AMP) levels of the fr mutant, a morphologically aberrant strain of Neurospora crassa, are reduced 2- to 5-fold. By taking advantage of the differences in phenotype of fr in liquid and agar cultures and the positive response of fr grown on solid support to exogenous theophylline, a relationship between the degree of morphological abnormality and intracellular cyclic AMP levels of the mutant is observed. Progressive restoration of the fr phenotype toward a normal state is paralleled by increases in cyclic nucleotide content. Striking differences in the sedimentation and thermal characteristics of the fr and wild-type adenylate cyclases [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] are observed. Approximately 50% of the normal activity sediments at 105,000 X g compared to 5% of the mutant enzyme. In addition, the overall stability of the fr adenylate cyclase is significantly decreased and its rate of inactivation at 37 degrees in the absence of substrate is 10-fold greater than the wild-type adenylate cyclase. Arrhenius plots also indicated that the Q10 (increase in rate per 10 degrees temperature increase) and the temperature of maximal activity of the fr enzyme are reduced. Supplementation of fr agar cultures with linolenic acid results in an elevated cyclic AMP content and a wild-type-like morphology similar to that obtained with inhibitors of phosphodiesterase (3':5'-cyclic AMP 5'-nucleotidohydrolase, EC 3.1.4.17). An increased thermostability of the fr adenylate cyclase occurs on linolenate enrichment of the mutant. It is concluded that the cyclic AMP deficiency is at least partially responsible for the fr phenotype and that this reduction results from a membrane defect that affects adenylate cyclase function.
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PMID:Adenosine 3':5'-cyclic monophosphate deficiency in Neurospora crassa. 18 53


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