Showing papers on "Cyclase published in 1979"

Activation of adenylate cyclase by choleragen.

Abstract: PERSPECTIVES AND SUMMARY S82 ROLE OF GANGLIOSIDE GMI AS THE CELL SURFACE RECEPTOR FOR CHOLERAGEN ......... ...... . . . ....... . ......... ...... .... 583 Interaction of Gangliosides and Their Oligosaccharides with Choleragen and Its Subunits 584 Activation of Adenylate Cyclase by Choleragen in Intact Cells 585 Activation of Adenylate Cyclase by Choleragen in Cell-Free Systems . 587 REQUIREMENTS FOR DEMONSTRATION OF ADENYLATE CYCLASE ACTIVATION BY CHOLERAGEN IN CELL-FREE SySTEMS ....... . .. 588 NAD !i88 GTP 588 Calcium-Dependent Regulatory Protein 589 ROLE OF NAD AS SUBSTRATE IN REACTIONS CATALYZED BY CHOLERAGEN 589 Mechanism of Action of NAD-Dependent Pseudomonas EXotoxin A and Diphtheria Toxin .... . . . ....... ..... ......... 590 NAD Glycohydrolase and ADP-Ribosyltransferase Activities oj Choleragen 591 Effects ofCholeragen on GTP-Binding Protein and GTPase Activity 592 SIMILARITIES BETWEEN CHOLERAGEN AND ESCHERICHIA COLI HEAT-LABILE ENTEROTOXIN 593 AN ADP-RIBOSYLTRANSFERASE FROM TURKEY ERYTHROCYTES WITH CHOLERAGEN-LIKE ACTIVITy .... . . . . . . . . . . . 595

The Biosynthesis and Metabolism of Prostaglandins

Abstract: With improved techniques for isolation and identification of materials, thromboxane (TXA2) and prostacyclin (PGI2) derivatives are now recognized as more abundant in some tissues and more potent than PGE2 and PGF2alpha. The rapid appearance and disappearance of these autacoids can be regulated at many points along the enzymatic path. Two important features affecting the rate of overall prostaglandin formation are the availability of non-esterified substrate and the availability of hydroperoxide activator for the cyclooxygenase. The fate of the endoperoxide formed by this reaction then depends upon the different relative amounts of the synthases and dehydrogenases in each type of synthesizing cell. Important future developments will indicate ways in which the amounts of these enzyme activities are altered and the ways in which the prostaglandin receptors interact with cellular adenyl cyclase and adrenergic receptors.

beta-Adrenergic receptor agonists increase phospholipid methylation, membrane fluidity, and beta-adrenergic receptor-adenylate cyclase coupling

Abstract: The beta-adrenergic agonist L-isoproterenol stimulated the enzymic synthesis of phosphatidyl-N-monomethylethanolamine and phosphatidylcholine in rat reticulocyte ghosts containing the methyl donor S-adenosyl-L-methionine. The stimulation was stereospecific, dose-dependent, and inhibited by the beta-adrenergic agonist propranolol. The addition of GTP inside the resealed ghosts shifted the dose-response of phospholipid methylation by L-isoproterenol to the left by 2 orders of magnitude. Direct stimulation of adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] with sodium fluoride or cholera toxin did not increase the methylation of phospholipids. At a concentration of S-adenosyl-L-methionine that stimulates synthesis of phosphatidyl-N-monomethylethanolamine, the activity of isoproterenol-sensitive adenylate cyclase was increased 2-fold without changes in the basal activity of adenylate cyclase and the number of beta-adrenergic receptors. The increase of phospholipid methylation by L-isoproterenol decreased membrane viscosity and increased translocation of methylated lipids. These findings indicate that enhancement of phospholipid methylation by L-isoproterenol decreases membrane microviscosity and thus increases lateral movement of the beta-adrenergic receptors and coupling with adenylate cyclase.

Dopamine inhibits adenylate cyclase in human prolactin-secreting pituitary adenomas

Abstract: THERE is evidence for the existence of multiple forms of dopamine (DA) receptors1–3. In particular, some of these receptors are coupled to adenylate cyclase (DA-stimulating enzyme activity), whereas others seem to be independent3–5. These two classes of receptors, which have been designated D-1 and D-2, respectively, would also have different pharmacological properties3. Pituitary mammotroph DA receptors regulating prolactin (PRL) secretion are considered to be typical D-2 receptors. A DA-stimulated adenylate cyclase has not been detected in normal anterior pituitaries3,5,6; furthermore, several studies indicate that cyclic AMP is stimulatory and not inhibitory to PRL secretion7,8. We report here that in homogenates of human PRL adenomas, in which dopaminergic agonists act as inhibitors of PRL secretion, basal adenylate cyclase is inhibited by DA. The DA receptors mediating this inhibition have the same pharmacological properties as those regulating PRL secretion.

The pharmacological specificity of beta-1 and beta-2 adrenergic receptors in rat heart and lung in vitro.

Abstract: The potency and selectivity of a variety of agonists and antagonists were determined for β-1 and β-2 adrenergic receptors on membranes prepared from rat ventricular muscle and lung. Activation or inhibition of β-adrenergic receptor stimulated adenylate cyclase activity and inhibition of specific (125I)-iodohydroxybenzylpindolol binding were used as in vitro measurements of receptor occupancy. With both assays the relative potencies of isoproterenol, epinephrine and norepinephrine with cardiac membranes was approximately 1:10:10, indicating a population of mainly β-1 adrenergic receptors. With membranes from lung the order of potency of these compounds was approximately 1:10:100, indicating a population mainly of β-2 adrenergic receptors. Several drugs previously reported to be β-2 selective agonists (salbutamol, soterenol, salmefamol, zinterol and fenoterol) activated adenylate cyclase in the lung but not in the heart. These compounds turned out to be partial agonists and isoproterenol-stimulated adenylate cyclase activity was inhibited by them in both tissues. Several compounds previously reported to be either β-1 (dobutamine) or β-2 (terbutaline and metaproterenol) selective agonists had similar potencies for stimulation of adenylate cyclase from both tissues. A series of compounds reported to be β-1 selective antagonists were also investigated. Metoprolol and practolol were 10-fold, and atenolol was 3-fold more potent in the heart than the lung. Butoxamhe, a β-2 antagonist, was 2-4 fold more potent in the lung than the heart, while H35/25 showed no specificity. The ability of antagonists to inhibit [125I]-iodohydroxybenzylpindolol binding to membranes prepared from the heart and lung agreed well with their effects on adenylate cyclase. The β-2 selective agonists zinterol and salmefamol also showed a 10-50 fold greater potency in inhibiting [125I]-iodohydroxybenzylpindolol binding in the lung than the heart. However salbutamol, soterenol and fenoterol, which selectively activated adenylate cyclase in the lung, inhibited [125I]-iodohydroxybenzylpindolol binding in the two tissues with equal potency. This apparent discrepancy appears to be due to the fact that these drugs which are partial agonists in the lung are competitive antagonists in the heart, and that the Ki values in the heart are very similar to the K act values in the lung.

Calmodulin activation of adenylate cyclase in pancreatic islets

Abstract: Pancreatic islets contain calmodulin. The protein binds to a particulate fraction derived from the islets and stimulates adenylate cyclase activity in this subcellular fraction, both phenomena being activated by ionized calcium. A calcium-dependent stimulation of adenylate cyclase by endogenous calmodulin may contribute to the accumulation of adenosine 3',5'-monophosphate evoked by insulin releasing agents in the islet cells.

Regulation of adenylate cyclase by adenosine.

Abstract: Adenosine may well be as important in the regulation of adenylate cyclase as hormones. Sattin and Rall first demonstrated in 1970 that adenosine was a potent stimulator of adenylate cyclase in the brain. However, adenosine is an equally potent inhibitor of adenylate cyclase in other cells such as adipocytes. The concentration of adenosine required for this regulation of adenylate cyclase is in the nanomolar range (10 to 100 nm). Both the inhibitory and stimulatory effects of low concentrations of adenosine on adenylate cyclase are antagonized by methylxanthines. This antagonism of adenosine action may account for all or part of the effects of methyl xanthines on cyclic AMP levels in many tissues. Adenosine appears to be a particularly important endogenous regulator of adenylate cyclase in brain, smooth muscle and fat cells. Under conditions in which intracellular AMP rises, adenosine formation and release is accelerated. In addition to its direct effects on adenylate cyclase, adenosine (at higher concentrations approaching millimolar) exerts multiple effects on cellular metabolism as a result of its intracellular metabolism and especially conversion to nucleotides.

Coupling of opiate receptors to adenylate cyclase: requirement for Na+ and GTP

Abstract: Inhibition of the adenylate cyclase activity in homogenates of mouse neuroblastoma-glioma hybrid cells (NG108-15) by the opioid peptide [D-Ala2,Met5]enkephalin amide (AMEA) requires the presence of Na+ and GTP. In this process, the selectivity for monovalent cations is Na+ greater than or equal Li+ greater than K+ greater than choline+; ITP will replace GTP but ATP, UTP, or CTP will not. The apparent Km for Na+ is 20 mM and for GTP it is 1 microM. Under saturating Na+ and GTP conditions, the apparent Ki for AMEA-directed inhibition is 20 nM for basal and 100 nM for prostaglandin E1-activated adenylate cyclase activity. For both cyclase activities, maximal inhibition is only partial (i.e., approximately 55% of control in each case). In intact viable NG108-15 cells, the decrease in basal and prostaglandin E1-stimulated intracellular cyclic AMP concentrations by AMEA is also dependent upon extracellular Na+. The enkephalin-directed reductions in cyclic AMP concentrations are at least 75%. The specificity of the monovalent cation requirement for enkephalin action on intact cells is the same as for enkephalin regulation of homogenate adenylate cyclase activity. Based on these data, a model is presented in which the transfer of information from opiate receptors to adenylate cyclase requires active separate membrane components, which correspond to the sites of action of Na+ and GTP in this process.

The Adenylate cyclase-cyclic AMP system in islets of langerhans and its role in the control of insulin release

Abstract: The mechanisms by which glucose and other stimulators control the rate of insulin release from the /3-cells of islets of Langerhans is still far from understood. In general terms, it is assumed that a receptor for a glucose metabolite is the initiator of interactions leading to increased insulin release. These interactions may involve cyclic AMP, Ca ++ , cytosol components, microfilaments and microtubules, the insulin containing granule membranes and the Ncell plasma membrane, and lead to fusion of granule membranes with the plasma membrane and the release of insulin from the cell. This short review concentrates on the adenylate cyclase system including the enzyme, its product cyclic AMP, cyclic nucleotide phosphodiesterases which break down cyclic AMP, effector systems for cyclic AMP such as protein kinases and phosphoprotein phosphatases. Inevitably, the interaction with glucose will be an important theme in the discussion because (a) cyclic AMP \"potentiates\" the insulin-releasing capability of glucose and (b) glucose increases the intracellular concentration of cyclic AMP in islets of Langerhans. Knowledge that cyclic AMP enhances the/3-cell response to glucose came from the demonstration that glucagon increased insulin secretion and that the response to glucagon was greater during hyperglycaemia that at normal glucose levels [1-4]. By analogy with the effect of glucagon in stimulating adenylate cyclase in liver, it was suggested that cyclic AMP was involved in the mediation of these effects. Since that time, many reports have confirmed the effects of glucagon on insulin release, and the similarities of action between those agents which raise intracellular cyclic AMP levels, such as glucagon, /3-adrenergic agents, phosphodiesterase inhibitors (papaverine, methylxanthines) and cholera toxin, and the effects of exogenous cyclic AMP or dibutyryl cyclic AMP have been noted [5-14]. It is clear from these reports that when the cyclic AMP concentration of the/3-cell is raised, by whatever means, the insulin response to glucose stimulation is enhanced. It is also clear that the effects of cyclic AMP are greatest at high glucose concentrations, whereas at low, non-stimulatory glucose concentrations the effects of cyclic AMP are usually small.

Activation of adenylate cyclase by vanadate.

Abstract: VANADATE has been identified as a potent inhibitor of (Na,K)ATPases1,2. The (Na,K)ATPase of dog kidney is inhibited 50% by 40 nM sodium orthovanadate (Na2VO4) in optimal conditions (28 mM Mg2+)2. As vanadate was originally identified as an inhibitor in muscle-derived ATP preparations2 and in equine and rabbit skeletal muscles, it has been suggested as endogenous regulator of (Na,K)ATPase2–4. We have tried to use vanadate as a tool to inhibit ATPases in adenylate cyclase studies when working at low ATP concentrations. We report here that vanadate causes marked stimulation of adenylate cyclase activity in membranes isolated from rat fat cells and produces maximal activation even in the absence of hormones.

Resolution of adenylate cyclase sensitive and insensitive to Ca2+ and calcium-dependent regulatory protein (CDR) by CDR-Sepharose affinity chromatography

Abstract: Partially purified adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] from bovine brain cortex was fractionated into two separate forms by calcium-dependent regulatory protein (CDR)-Sepharose affinity chromatography. The major form of the enzyme, comprising approximately 80% of the applied activity, did not bind to the affinity column in the presence of Ca2+ and was insensitive to the CDR. Approximately 20% of adenylate cyclase activity was absorbed to CDR-Sepharose in the presence of Ca2+. This activity was stimulated by Ca2+ and CDR. This study directly demonstrates that brain cortex contains Ca2+-CDR-sensitive and -insensitive forms of adenylate cyclase and indicates that CDR-Sepharose may be a useful tool for purification of adenylate cyclase. The Ca2+ -stimulated adenylate cyclase was purified at least 55-fold with a 13% yield.

Evidence that cyclic GMP regulates membrane potential in rod photoreceptors

Abstract: CYCLIC NUCLEOTIDES have been proposed as mediators of the decreased Na+ permeability caused by illumination of vertebrate photoreceptors1,2. We show here that cyclic GMP depolarises the rod outer segment (ROS) approximately to the Na+ equilibrium potential within milliseconds after being injected intracellularly, that previous light adaptation antagonises this depolarisation, and that the injection of cyclic GMP without illumination initiates a repolarisation after a time lag which is longer than that following a light flash and proportional to the injection time. The results are interpreted in terms of a model in which cyclic GMP levels are controlled by the resultant of cyclase and phosphodiesterase (PDE) velocities. The PDE velocity of hydrolysis in the dark is increased by prior light adaptation in the presence of increased substrate (cyclic GMP). The data suggest that cyclic GMP is an important factor in the regulation of the membrane potential of the ROS.

Effects of Arachidonic Acid and Its Metabolites on Antigen-Induced Histamine Release from Human Basophils in Vitro

Abstract: We have studied the role of arachidonic acid (AA) metabolism in immediate hypersensitivity reactions. The AA analog, eicosa-5,8,11,14-tetraynoic acid (ETYA), which inhibits both the cyclo-oxygenase and lipoxygenase pathways of AA metabolism, was found to cause a dose-dependent inhibition of antigen-induced histamine release from human basophils; complete inhibition occurred at 10 -4 M. In contrast, nonsteroidal anti-inflammatory drugs (NSAID) including indomethacin (10 -9 to 10 -6 M), meclofenamic acid (10 -6 to 10 -4 M), and aspirin (10 -5 to 10 -3 M), which block the cyclo-oxygenase pathway of AA metabolism, enhanced mediator release. AA (10 -7 to 10 -5 M) also enhanced basophil mediator release, an effect that was not affected by preincubation with indomethacin. Other fatty acids including linoleic, linolenic, and stearic acids were without effect. Indomethacin (and the other NSAID studied) reversed the inhibition caused by the AA metabolite, prostaglandin (PG) E 2 , in a dose-dependent fashion. This reversal of inhibition was not unique to PG, but was observed with all inhibitors of histamine release studied that act via activation of adenylate cyclase including isoproterenol and the H2 agonist, dimaprit. NSAID, however, did not reverse the inhibition caused by drugs that do not act on adenylate cyclase, i.e., dibutyryl cAMP, isobutylmethylxanthine, and 2-deoxyglucose. In the leukocyte preparations studied, the NSAID did not block the increase in the intracellular cAMP levels caused by agonists that act via adenylate cyclase. It appears that a product(s) of AA metabolism, generated via the lipoxygenase pathway, is involved in IgE-mediated histamine release and that the same or a different product(s) modulates the control mechanisms that are linked to adenylate cyclase.

Effect of gangliosides on adenylate cyclase activity in rat cerebral cortical membranes.

Abstract: The addition of 50 µM mixed brain gangliosides to membrane preparations from rat cerebral cortex caused a 50-95% increase in the basal adenylate cyclase activity. The activation was rapid at all temperatures and was relatively irreversible, as evidenced by the fact that repeated washing of the membrane after exposure to gangliosides failed to restore adenylate cyclase activity to its basal level. The V max of the enzyme was increased with no apparent change in the K m for the substrate, ATP. The expression of activation of adenylate cyclase by gangliosides showed a marked temperature dependence, with maximal effects seem in the 30-40° range and no activation seen at either 10° or 50°. The activation of adenylate cyclase by brain gangliosides was additive with respect to activation by NaF and detergents, but was not additive with respect to activation by GppNHp or lysolecithin. The activation of the enzyme by gangliosides was unaffected by the presence of calcium ions, calcium-dependent activator protein, or EGTA. The addition of brain gangliosides to the membranes caused a consistent restoration of the responsiveness of the adenylate cyclase system to activation by β-adrenergic agents, an effect similar in magnitude to that observed by addition of GppNHp to the membrane preparations.

Role of intracellular Ca2+ sequestration in beta-adrenergic relaxation of a smooth muscle.

Abstract: Various mechanisms have been proposed for β-adrenergically mediated relaxation of smooth muscle. All theories suggest the involvement of cyclic AMP as a second messenger: β-agonists stimulate adenylate cyclase which converts ATP to cyclic AMP1 and protein kinase, activated by cyclic AMP, is then thought to catalyse a protein phosphorylation that leads to a reduction in free Ca2+, thus effecting relaxation1. How this last step is accomplished is much debated, but the following possibilities are currently considered as the mechanisms responsible for cyclic AMP-induced reduction of cytoplasmic Ca2+: activation of a Ca2+-ATPase in the plasma and/or sarcoplasmic reticulum membranes which lowers cytoplasmic [Ca2+] in a direct manner or stimulation of (Na+–K+)ATPase in the cell membrane which may indirectly effect Ca2+ extrusion2–8. Among the hypotheses suggested, those of Ca2+ sequestration by the sarcoplasmic reticulum and of Ca2+ extrusion across the cell membrane are consistent with each other if it is assumed that both processes are effected by a cyclic AMP-sensitive Ca2+-ATPase. However, quite a different mechanism is implied by involving the Na+–K2+ pump and Na+–Ca2+ exchange carrier. In this report, we present evidence that suggests intracellular Ca2+ sequestration is the mechanism involved.

Mutations in cyclic AMP-dependent protein kinase and corticotropin (ACTH)-sensitive adenylate cyclase affect adrenal steroidogenesis.

Abstract: Two groups of mutant clones were isolated from YI adrenocortical tumor cells. One group, Y1(Kin), exhibited altered cytosolic cyclic AMP-dependent protein kinase activity; the second group, Y1(Cyc), exhibited diminished corticotropin-responsive adenylate cyclase activity. Steroidogenic responses to corticotropin and cyclic nucleotides closely paralleled cyclic AMP-dependent protein kinase activity in the Y1(Kin) mutants. In Y1(Cyc) mutants, corticotropin had little effect on steroidogenesis, whereas cyclic nucleotides were fully active. These data imply that adenylate cyclase and cyclic AMP-dependent protein kinase are obligatory components of the corticotropin-stimulated steroidogenic pathway.

Isoproterenol-Induced Cardiac Hypertrophy: Modifications in Characteristics of β-Adrenergic Receptor, Adenylate Cyclase, and Ventricular Contraction

Abstract: Several modifications in characteristics of the β- adrenergic receptor-adenylate cyclase system were observed in the cardiac hypertrophy of the rat produced by chronic isoproterenol treatment. These included 1) a decreased number of β- adrenergic receptors without changes in affinity when assayed by (-).[3H]dihydroalprenolol binding, 2) a decreased ability of 5′-guanylylimidodiphosphate to inhibit displacement by isoproterenol of the bound (-)-[3H]dihydroalprenolol, 3) a decreased sensitivity and magnitude of adenylate cyclase in heart homogenates to isoproterenol stimulation coupled with decreases in basal and NaF-stimulated enzyme activity, 4) a decreased responsiveness of the incubated heart minces to isoproterenol stimulation with respect to cAMP formation, and 5) a decreased contractile force development in ventricular strips in response to isoproterenol. All of the above alterations associated with the isoproterenol-induced cardiomegaly disappeared upon regressio n of hypertrophy. Light and electron...