Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01275 (glucagon)
 26,492 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Adenylate cyclase (EC 4.6.1.1) activity in mouse liver plasma membranes is increased fivefold when animals are pretreated with cholera toxin. The increase in activity is detectable within 20 min of an intravenous injection of the toxin. The response of the control and cholera-toxin-activated adenylate cyclase to hormones, GTP, and NaF is complex. GTP causes the same fold stimulation of control and toxin-activated cyclase, but glucagon and NaF remain the most potent activators of liver adenylate cyclase irrespective of whether the enzyme is activated by cholera toxin. Determination of kinetic parameters of adenylate cyclase indicates that cholera toxin, hormones, and NaF do not change the affinity of the enzyme for ATP-Mg nor do they alter the Ka for free Mg2+. High concentrations of Mg2+ inhibit adenylate cyclase that is stimulated by either cholera toxin, glucagon, or NaF. These same Mg2+ concentrations have no effect on the basal activity of the enzyme or its activity in the presence of GTP.
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PMID:Cholera toxin and adenylate cyclase: properties of the activated enzyme in liver plasma membranes. 100 Mar 62

Incubation of rat fat pad membranes with 5-guanylyliminodiphosphonate [Gpp-(NH)p] and 5-guanylylmethylenediphosphonate [Gpp(CH2)p], but not GTP (with or without hormones), at 24 degrees or 30 degrees (but not at 4 degrees) greatly stimulates adenylate cyclase activity [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] measured after thoroughly washing the membranes. The rate of activation is relatively slow, even with very high (and saturating) concentrations of the analogs. Binding alone appears to be insufficient for activation. Hormones (catecholamines, glucagon) increase the rate but not the extent of activation, even when saturating analog concentrations are used. The dependence on analog concentration (apparent Km) varies with the time of incubation. GTP and very high concentrations of ATP inhibit the activation by Gpp(NH)p, but this effect is dependent on the length of incubation and can be overcome with time. The activated state is not reversed upon incubation of the washed membranes at 30 degrees, even in the presence of GTP, or by solubilization with nonionic detergents. Also, Gpp(NH)p can directly stimulate the control, solubilized enzyme. The activated state of the solubilized enzyme persists upon specific adsorption to and subsequent elution from an organomercurial-agarose column. It is suggested that after forming reversible Michaelis complexes of relatively low affinity, these analogs may react irreversibly with the GTP regulatory site of the enzyme, perhaps forming p(NH)p- and p(CH2)p-covalent enzyme intermediates which capture the activated state of the enzyme. GTP, after binding, may normally activate the enzyme by forming a "labile" pyrophosphoryl enzyme intermediate, and hormone receptors may function to increase the rate of formation (and thus concentration) of this active state of the enzyme.
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PMID:Activation of adenylate cyclase by phosphoramidate and phosphonate analogs of GTP: possible role of covalent enzyme-substrate intermediates in the mechanism of hormonal activation. 105 66

The effects of various nucleosides and nucleotides upon glucagon secretion from the isolated perfused rat pancreas were studied. Increasing glucagon secretion was found with increasing concentrations of exogenous cyclic AMP (2 X 10(-4) M, 2 X 10(-3) M and 1 X 10(-2) M). Stimulation of alpha cell secretion was also found with 2 X 10(-3) M 2'AMP, 3'AMP, 5'AMP, ADP, Adenosine, NADP, and NADPH. One X 10(-3) M cyclic GMP elicited significant glucagon secretion. The pattern of glucagon release was similar in all cases with peak secretion occurring during the 30- to 90-s time period following initiation of the stimulus. No significant increase of glucagon secretion was found in response to ATP, guanosine, 2'GMP, 3'GMP, 5'GMP, GTP, xanthosine, inosine, adenine, xanthine, thymidine, cytidine, ribose, nicotinamide, and uric acid. On the basis of the above results, the structural requirement for stimulation of glucagon secretion appears to be adenine linked to ribose, with phosphate groups being unnecessary. The conclusion of this study is that a new class of compounds capable of stimulating glucagon secretion has been identified, and important questions are thus raised about the mechanism of the action of exogenous cyclic AMP.
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PMID:Nucleotide and nucleoside stimulation of glucagon secretion. 110 53

Previous studies have shown that guanine nucleotides, acting at a site termed nucleotide regulatory site, are required for activation of hepatic adenylate cyclase and that glucagon facilitates this process. This study shows that only guanine nucleotides containing triphosphate groups at the 5' position of ribose (or 3'-deoxyribose) are capable of activating the enzyme. The terminal phosphate is not utilized in the activation process since 5'-guanylylimidodiphosphate (Gpp(NH)p and 5'-guanylyl methylenediphosphonate, analogues of GTP that are not utilized in transferase or hydrolase reactions, stimulate enzyme activity. The nucleotides bind in their free form at the regulatory site; chelation by magnesium ion shifts the apparent concentration dependence for activation by Gpp(nh)p. GDP inhibits competitively Gpp(NH)p-stimulated activity and inhibits basal activity and activities stimulated by glucagon. Activation of the enzyme by Gpp(NH)p is a slow process; the length of the lag time increases as an inverse function of nucleotide concentration and is as long as 4 min before onset of increased enzyme activity. Following pretreatment with Gpp(NH)p and extensive washing of hepatic membranes, the enzyme displays immediate increases in activity with rates that are a function of the nucleotide concentration during pretreatment; the rates remain constant for at least 6 min despite the absence of Gpp(NH)p in the medium. Studies with labeled Gpp(NH)p show that the intact nucleotide remains firmly bound to the membranes after extensive washing, suggesting that the persistence of adenylate cyclase activity may be related to slow dissociation of the nucleotide from the regulatory site. Addition of 1 nM glucagon, a submaximal concentration, does not abolish the lag phase of Gpp(NH)p activation even at saturating concentration of the nucleotide (1 muM or higher). The maximal steady state rate is achieved under these conditions. Addition of 2 muM glucagon, a saturating hormone concentration, does not alter the steady state rate but abolishes the lag phase of Gpp(NH)p activation. The transient kinetics of Gpp(NH)p activation and the effects of glucagon thereon are discussed in terms of a three state model in which the guanine nucleotide induces the formation of an intermediate transition state that displays no increase in enzyme activity over the basal state and which slowly isomerizes to a high activity state of the adenylate cyclase system; glucagon acts by accelerating the rate of isomerization.
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PMID:The hepatic adenylate cyclase system. I. Evidence for transition states and structural requirements for guanine nucloetide activiation. 112 49

The epinephrine sensitivity in vitro of the adenylate cyclase system in liver plasma membranes from adrenalectomized rats was increased by the addition of 1 to 100 muM GTP or GDP in the incubation medium. Basal and glucagon-stimulated cyclase activities were also enhanced by GTP and GDP. These effects occurred even in the absence of an ATP-regenerating system. They were mimicked by 5'-guanyl diphosphonate and a series of guanyl derivatives, indicating that the structural requirement for the GTP action is not very stringent. Guanyl nucleotides did not increase the affinity of the adenylate cyclase system for the activating hormones, nor did they protect the enzyme activity against denaturation. Their synergic effect was due to an enhancement of the affinity of the enzyme for the substrate MgATP and also to an increase of the maximal velocity of the reaction. It is proposed that the guanyl nucleotides act directly and primarily upon the catalytic component of the cyclase system, independently of their effects on the binding of the activating hormones to liver plasma membrane. Since the activating effects of epinephrine and glucagon are similar in the presence of GTP, but not in its absence, it is suggested that the lower efficiency of epinephrine under normal conditions is not due to intrinsic membrane characteristics, but rather, to superimposed extraneous modulations.
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PMID:The epinephrine-sensitive adenylate cyclase of rat liver plasma membranes. Role of guanyl nucleotides. 114 Dec 21

Glucagon and adrenaline exert their action upon the liver via the cyclic AMP synthetizing system located in the plasma membrane. The enzyme adenylate cyclase is further regulated by guanyl nucleotides. It has been recently shown that the rat liver plasma membrane system could respond to GTP by simultaneous increase in the cyclase activity in response to glucagon and by the dissociation of this hormone from its binding sites (1). Unambiguous relationship between the activating effect of GTP upon the cyclase and its action upon glucagon binding has not been determined yet (2). This problem was approached using the in vitro action of epinephrine as a model. When 1 to 100 muM GTP or DGP were added to rat liver plasma membranes isolated from adrenalectomized animals, they increased markedly the response of the cyclase system to epinephrine. These effects could be observed in the absence of an ATP-regenerating system and were mimicked by 5'-guanylyl diphosphonate; GTP and GDP were the most active compounds followed by ITP, CTP and by a series of guanyl derivatives. UTP, as well as guanosine, GMP, cyclic GMP and ppGpp were inactive. Guanyl nucleotides did not increase the affinity of the cyclase system for the activating hormones, but enhanced the affinity for ATP-Mg and also the Vmax of the reaction. Finally, GTP, ATP, CTP, UTP but not GDP displaced epinephrine bound to plasma membranes by a mere chelation phenomenon. It is concluded that 1) guanyl nucleotides do not act primarily by influencing the binding of hormones to the membranes; 2) they act directly upon the catalytic subunit of the cyclase; 3) the low concentrations of GTP required for its action strongly suggest that this nucleotide plays a role in the physiological regulation of the intrahepatic cyclic AMP level.
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PMID:[Role of guanidylic nucleotides in the adenylate cyclase activity of the rat liver]. 120 15

Glucagon, a peptide hormone synthesized and secreted by alpha islet cells, regulates glucose homeostasis by several mechanisms. Using [gamma 32P]8N3GTP, a proven photoaffinity probe for GTP, a specific nucleotide binding site on human glucagon was detected that showed preference for GTP. Half-maximal saturation of photoinsertion into the polypeptide hormone was at 8-12 microM with either [alpha 32P]8N3GTP or [gamma 32P]8N3GTP. GTP protected photolabeling with an apparent kd of 15 microM, whereas ATP was less effective as a protector, exhibiting an apparent kd of about 30 microM. Maximal protection by GTP and ATP was over 90%. UTP, CTP, GDP, ADP, GMP, AMP, guanosine, adenosine, guanine, and adenine were much less effective protectors, indicating that binding is specific for purine nucleoside triphosphates, particularly GTP. Mg2+ at 150 microM enhanced photoinsertion (twofold), whereas at 2-10 mM, it inhibited photoinsertion. Both Ca2+ and Zn2+ at 0.2 mM decreased photoinsertion about 45%. Purification of chymotryptic and tryptic digests of photolabeled glucagon by reverse-phase high performance liquid chromatography (HPLC) revealed that the N-terminal peptide, HSQGTF, was the only peptide region covalently photomodified by [32P]8N3GTP. GTP, if present during photolysis, greatly reduced both photoinsertion into glucagon and the amount of radiolabeled peptide recovered on HPLC. The specificity of binding to the N-terminal region is suggestive of a physiological role for a glucagon-GTP complex in the mechanism of action of this hormone.
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PMID:Identification of the guanine binding domain peptide of the GTP-binding site of glucagon. 130 73

In Zajdela hepatoma cells (ZHC) the plasma membrane Ca2+ pump displayed no sensitivity to glucagon (19-29) (mini-glucagon), whereas in hepatocyte this metabolite of glucagon evoked a biphasic regulation of the Ca2+ pump system via a cholera toxin-sensitive G protein. Analysis of G protein subunits in ZHC membranes indicated the presence of cholera toxin-sensitive Gs alpha and G beta gamma proteins, whose functionality was manifested by GTP and NaF stimulation of adenylylcyclase activity, and pertussis toxin-catalyzed ADP-ribosylation of Gi alpha, respectively. However, immunoblotting experiments suggested a lower content in beta gamma subunits in ZHC as compared with hepatocyte plasma membranes. Complementation of ZHC or hepatocyte plasma membranes with purified beta gamma subunits from transducin (T beta gamma) caused inhibition of the basal activity of the Ca2+ pump at 10 and 300 ng/ml, respectively, and revealed (in ZHC) or increased (in hepatocytes) sensitivity of the system to mini-glucagon. After cholera toxin treatment of ZHC, T beta gamma no longer reconstituted the response of the Ca2+ pump to mini-glucagon, suggesting that the mechanism of beta gamma action is dependent on an association with the alpha subunit of a cholera toxin-sensitive G protein. It is concluded that G beta gamma subunits control both the basal activity of the plasma membrane Ca2+ pump and its inhibition by mini-glucagon.
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PMID:Role of G protein beta gamma subunits in the regulation of the plasma membrane Ca2+ pump. 131 Mar 15

Glucagon-like peptide-1 (7-36)amide (GLP-1 (7-36)amide) represents a physiologically important incretin in mammals including man. Receptors for GLP-1 (7-36)amide have been described in RINm5F cells. We have solubilized active GLP-1(7-36)amide receptors from RINm5F cell membranes utilizing the detergents octyl-beta-glucoside and CHAPS; Triton X-100 and Lubrol PX were ineffective. Binding of radiolabeled GLP-1(7-36)amide to the solubilized receptor was inhibited concentration-dependently by addition of unlabeled peptide. Scatchard analysis of binding data revealed a single class of binding sites with Kd = 0.26 +/- 0.03 nM and Bmax = 65.4 +/- 21.24 fmol/mg of protein for the membrane-bound receptor and Kd = 22.54 +/- 4.42 microM and Bmax = 3.9 +/- 0.79 pmol/mg of protein for the solubilized receptor. The binding of the radiolabel to the solubilized receptor was dependent both on the concentrations of mono- and divalent cations and the protein/detergent ratio in the incubation buffer. The membrane bound receptor is sensitive to guanine-nucleotides, however neither GTP-gamma-S nor GDP-beta-S affected binding of labeled peptide to solubilized receptor. These data indicate that the solubilized receptor may have lost association with its G-protein. In conclusion, the here presented protocol allows solubilization of the GLP-1(7-36)amide receptor in a functional state, and opens up the possibility for further molecular characterization of the receptor protein.
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PMID:Solubilization of active GLP-1 (7-36)amide receptors from RINm5F plasma membranes. 131 74

We studied the effect of bromolevamisole (BL) and other imidazo [2,1-b] thiazole derivatives--bromodexamisole (BD) and levamisole (LV)--on adenylate cyclase (AC) activity. BL and BD both inhibited forskolin-activated human thyroid AC, while LV had no effect. This inhibition was non-stereospecific and the IC50 values, as measured with 1 mM ATP and 40 microM forskolin, were 0.95 and 0.80 mM for BL and BD, respectively. In contrast, human thyroid alkaline phosphatase (ALP) inhibition was stereospecific, with IC50 values of 0.0012 mM for BL and 0.9 mM for BD. LV was a 10-fold weaker inhibitor of ALP than BL. These results show that ALP inhibition is not correlated with forskolin-activated AC inhibition. Furthermore, in the presence of a competitive inhibitor of GTP (0.1 mM guanosine 5'-O-(2-thiodiphosphate), BL retained its antagonizing effect on forskolin-activated AC which suggests a direct action on the catalytic subunit. The inhibition was of the mixed type, indicating a complex interaction between BL and AC. Glucagon-activated AC activity in rat liver membranes was also inhibited by BL, although to a slightly lesser degree than thyroid stimulating hormone (TSH)-activated AC from human thyroid for a given BL concentration. In cultured human thyroid cells, BL (0.25 mM) induced a potent decrease in cAMP accumulation after 2 hr of stimulation by TSH. Taken together, these results show that BL inhibits AC and that this inhibition is not organ-specific.
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PMID:Effect of bromolevamisole and other imidazo [2,1-b] thiazole derivatives on adenylate cyclase activity. 131 3


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