UNIPROT:P01275 - FACTA Search

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Query: UNIPROT:P01275 (glucagon)
26,492 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The changes in glucagon receptors of white adipocytes from cold-acclimated rats were investigated to know the metabolic role of glucagon in cold acclimation by establishing a glucagon radioreceptor assay system for isolated white adipocytes. Glucagon radioreceptor assay methodology The binding of 125I-labelled glucagon to isolated epididymal white adipocytes was linearly related to the number of cells (0.5-2.0 X 10(5) cells/ml) added in the medium. At a cell concentration higher than 3.0 X 10(5) cells/ml, the amount of specific binding failed to show the proportional relationship to the number of adipocytes. The effects of incubation temperature (4 degrees C, 25 degrees C and 37 degrees C) on the glucagon binding were investigated. Incubation at 25 degrees C was adopted in the present study because of the highest maximum binding and the longest steady state obtained. Preincubation at 25 degrees C for 15 min increased significantly the amount of specific binding. It was confirmed that bacitracin, polypeptide antibiotics, inhibited significantly the degradation of glucagon. The glucagon binding under these conditions was found to be saturable and reversible, validating a specific reaction for the glucagon receptor. When a Scatchard plot was constructed, the data was curvilinear with an upward concavity, indicating the presence of at least two classes of binding site with different fixed affinities or of negatively cooperative interactions between receptors. It was concluded that an appropriate condition for glucagon receptor assay of white adipocytes consists of cell concentration of 1 X 10(5) cells/ml, 15 minute-preincubation and 30 minute-reaction at 25 degrees C in the presence of bacitracin (1 mg/ml). Effect of cold acclimation on glucagon receptors of white adipocytes Cold acclimation decreased the size and increased the number of epididymal white adipocytes. Cold acclimation increased the number of glucagon receptors of white adipocytes; about 140% increase expressed as per cell, approximately 260% increase per unit of surface area and 210% increase per whole tissue. The affinity of binding sites was not changed. The increased binding sites could explain, at least partly, the enhanced metabolic response of cold-acclimated rats to glucagon.
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PMID:[Studies on adipocyte glucagon receptor assay--with special reference to the effect of cold acclimation on glucagon receptors of white adipocytes]. 299 Oct 98

In this study, we determined the ability of four N-terminally modified derivatives of glucagon, [3-Me-His1,Arg12]-, [Phe1,Arg12]-, [D-Ala4,Arg12]-, and [D-Phe4]glucagon, to compete with 125I-glucagon for binding sites specific for glucagon in hepatic plasma membranes and to activate the hepatic adenylate cyclase system, the second step involved in producing many of the physiological effects of glucagon. Relative to the native hormone, [3-Me-His1,Arg12]glucagon binds approximately twofold greater to hepatic plasma membranes but is fivefold less potent in the adenylate cyclase assay. [Phe1,Arg12]glucagon binds threefold weaker and is also approximately fivefold less potent in adenylate cyclase activity. In addition, both analogues are partial agonists with respect to adenylate cyclase. These results support the critical role of the N-terminal histidine residue in eliciting maximal transduction of the hormonal message. [D-Ala4,Arg12]glucagon and [D-Phe4]glucagon, analogues designed to examine the possible importance of a beta-bend conformation in the N-terminal region of glucagon for binding and biological activities, have binding potencies relative to glucagon of 31% and 69%, respectively. [D-Ala4,Arg12]glucagon is a partial agonist in the adenylate cyclase assay system having a fourfold reduction in potency, while the [D-Phe4] derivative is a full agonist essentially equipotent with the native hormone. These results do not necessarily support the role of an N-terminal beta-bend in glucagon receptor recognition. With respect to in vivo glycogenolysis activities, all of the analogues have previously been reported to be full agonists.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Receptor binding and adenylate cyclase activities of glucagon analogues modified in the N-terminal region. 301 Oct 69

Delta-Tetrahydrocannabinol (delta 9-THC), the principal psychoactive constituent of Cannabis sativa, was found to increase glucagon activation of liver plasma membrane adenylate cyclase. In the presence of 30 microM delta 9-THC, the EC50 for glucagon was decreased by 60% from 7.6nM to 3.1 nM. 11-OH-delta 9-THC, a psychoactive metabolite of delta 9-THC, also increased glucagon activation of adenylate cyclase while two cannabinoids without marihuana-like psychoactive potency, cannabinol and cannabidiol, did not. At 30 microM, delta 9-THC either slightly decreased or had no effect on the activation of adenylate cyclase by GTP, Gpp(NH)p, fluoride ion, forskolin or ATP alone. Delta 9-THC had no effect on the binding of [125I] glucagon to liver plasma membranes. Arrhenius plots demonstrated that delta 9-THC and 11-OH-delta 9-THC, but not CBD, decreased the activation energy above the break temperature. Therefore, delta 9-THC increased the coupling of the glucagon receptor to adenylate cyclase apparently by removing a constraint on receptor-Ns coupling.
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PMID:Effects of delta 9-tetrahydrocannabinol on glucagon receptor coupling to adenylate cyclase in rat liver plasma membranes. 301 62

The role of the Tyr10-Ser11-Lys12-Tyr13 region of glucagon in the binding interaction and activation of the glucagon receptor was investigated by means of the synthetic glucagon analogues [Phe13]glucagonamide, [Phe10]glucagonamide, [Phe10]glucagon, [Phe10,13]glucagon, [Pro11]glucagon, [Pro11,Gly12]glucagonamide, [Ala11]glucagon, and [Oac11-13]glucagonamide. These analogues were synthesized by solid-phase peptide synthesis on p-methylbenzhydrylamine or Merrifield resins with protected N alpha-tert-butyloxycarbonyl amino acids. Purification by dialysis, cation-exchange chromatography, gel filtration, and preparative reverse-phase high-performance liquid chromatography (HPLC) gave products that proved homogeneous by thin-layer chromatography and HPLC and on analysis by amino acid analysis, by sequencing, and by alpha-chymotryptic peptide mapping with HPLC. Biological activities were examined by measurement of the stimulation of liver plasma membrane adenylate cyclase and by specific displacement of [125I]glucagon from glucagon receptors. The results of these studies indicate that while the biological "message" region of glucagon is located elsewhere, the 10-13 region has multiple roles in the glucagon-glucagon receptor interaction: this region provides functional groups for direct binding interaction with the receptor, and this region interacts with the receptor in such a way as to allow the "transduction message" portion of glucagon to interact and activate the receptor.
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PMID:Importance of the 10-13 region of glucagon for its receptor interactions and activation of adenylate cyclase. 301 6

We have previously demonstrated that the glucagon receptor binds hormone to form a low affinity complex which, by a time- and temperature-dependent mechanism, is converted to a high affinity complex (Horwitz, E.M., Jenkins, W.T., Hoosein, N.M., and Gurd, R.S. (1985) J. Biol. Chem. 260, 9307-9315). In this report we have investigated the effects of agonist concentration, potency, and intrinsic activity on the characteristics of the two, interconvertible states of the glucagon receptor. As the glucagon concentration is increased from 0.02 to 0.50 nM, the initial velocity of binding increases. The conversion of a low affinity to a high affinity complex is the rate-limiting step in the overall binding reaction and approaches its maximal velocity as the hormone concentration exceeds 0.20 nM. At equilibrium, 87-90% of the hormone-receptor complexes are in the high affinity state at all hormone concentrations examined. [S-methyl-Met27]glucagon, a full agonist with reduced potency, binds to the two-state system in a manner analogous to that of native glucagon. The binding of N alpha-biotinyl-N epsilon-acetimidoglucagon, a partial agonist with reduced potency, effects a two-state system where the high affinity state accounts for only 35% of the total hormone-receptor complexes at equilibrium. We conclude that the formation of the high affinity complex is the rate-limiting step involved in glucagon binding; reduction in binding potency with full agonism is due to a reduction in the affinity of the ligand for the unoccupied receptor and not to an alteration of the interconversion of the two states, and decreased intrinsic activity is due to a quantitative decrease in conversion of the low to high affinity state.
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PMID:Partial agonism in the glucagon receptor system is a consequence of the two-state rat hepatic receptor. 302 41

125I-Glucagon binding to rat liver plasma membranes was composed of high- and low-affinity components. N-Ethylmaleimide (NEM) and several other alkylating agents induced a dose-dependent loss of high-affinity sites. This diminished the apparent affinity of glucagon receptors for hormone without decreasing the binding capacity of membranes. Solubilized hormone-receptor complexes were fractionated as high molecular weight (Kav = 0.16) and low molecular weight (Kav = 0.46) species by gel filtration chromatography; NEM or guanosine 5'-triphosphate (GTP) diminished the fraction of high molecular weight complexes, suggesting that NEM uncouples glucagon receptor-N-protein complexes. Exposure of intact hepatocytes to the impermeable alkylating reagent p-(chloromercuri)benzenesulfonic acid failed to diminish the affinity of glucagon receptors on subsequently isolated plasma membranes, indicating that the thiol that affects receptor affinity is on the cytoplasmic side of the membrane. Hormone binding to plasma membranes was altered by NEM even after receptors were uncoupled from N proteins by GTP. These data suggest that a sensitive thiol group that affects hormone binding resides in the glucagon receptor, which may be a transmembrane protein. Alkylated membranes were fused with wild-type or cyc- S49 lymphoma cells to determine how alkylation affects the various components of the glucagon-adenylyl cyclase system. Stimulation of adenylyl cyclase with fluoride, guanylyl 5'-imidodiphosphate, glucagon, or isoproterenol was observed after fusion of cyc- S49 cells [which lack the stimulatory, guanine nucleotide binding, regulatory protein of adenylyl cyclase (Ns)] with liver membranes alkylated with 1.5 mM NEM.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:N-ethylmaleimide uncouples the glucagon receptor from the regulatory component of adenylyl cyclase. 302 2

The beta-adrenergic and glucagon receptor-binding capacities in rat livers from 6-27 months of age were measured to investigate the mechanism of a previously observed rise in beta-adrenergic stimulated adenylate cyclase with increasing age. There was no concomitant increase in glucagon-stimulated adenylate cyclase. In the present study neither glucagon-binding capacity nor glucagon-stimulated adenylate cyclase changed with age. In contrast, the beta-adrenergic receptor capacity, measured in the same membranes by [125I]iodopindolol binding, increased nearly 3-fold from 6.6 +/- 0.6 fmol/mg at 6 months to 19.1 +/- 3.3 fmol/mg at 18-19 months. The increase was directly proportional to the maximum isoproterenol-stimulated adenylate cyclase activity in livers of rats up to 19 months of age. By 24-27 months the binding capacity had increased to 24.9 +/- 3.3 fmol/mg, but there was no further increase in adenylate cyclase activity. Thus, there appeared to be a beta-receptor-adenylate cyclase uncoupling in livers from the senescent animals (25-27 months). The defect could not be demonstrated by studies examining isoproterenol competition of [125I]iodopindolol from agonist-induced high affinity sites on the membranes, a procedure that examines receptor-Ns protein coupling. Activation of adenylate cyclase by the nonhormonal stimulators F- and forskolin did not change with age, indicating that the catalytic unit was not a limiting factor. Since the relationship between the glucagon receptor and adenylate cyclase also remained unaltered, the uncoupling apparently lies in an alteration of the interaction between the beta-adrenergic receptor and the guanine nucleotide-sensitive Ns protein.
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PMID:Beta-adrenergic receptors, glucagon receptors, and their relationship to adenylate cyclase in rat liver during aging. 303 Jul 5

We studied the functional properties of hepatic glucagon receptors during rat development. Glucagon binding to liver membranes and isolated hepatocytes was significantly less in foetuses and weaning rats than in adult animals, reflecting changes in the number of receptors rather than any change in receptor affinity for the hormone. After correcting for the smaller surface area of foetal hepatocytes, glucagon receptor concentrations were still lower in foetuses than in adult rats. The time courses of glucagon association to liver membranes and of glucagon receptor degradation of prenatal and postnatal rats were similar, while inactivation of glucagon by liver membranes was significantly lower in foetal than in adult rats. Also, glucagon-stimulated production of cAMP was smaller in younger rats. These findings suggest that, according to several criteria, glucagon receptors in the foetus are functionally normal and their delayed development may be important for the metabolic processes occurring during the perinatal age.
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PMID:Characterization of glucagon receptors in liver membranes and isolated hepatocytes during rat ontogenic development. 303 Aug 50

In order to determine the role of glucagon in cold acclimation, the changes in glucagon receptor were investigated in white adipocytes from cold-acclimated rats by establishing a glucagon radioreceptor assay method for isolated white adipocytes. The following conditions were found to be appropriate for specific glucagon receptor binding assay; cell concentration of about 1 X 10(5) cells/ml, 15 min preincubation with glucagon and 30 min-reaction at 25 degrees C in the presence of bacitracin (1 mg/ml). Cold acclimation decreased the size and increased the number of epididymal white adipocytes. Cold acclimation increased the number of glucagon receptors of white adipocytes, resulting in 140% in terms of unit cell, and 260% increase per unit surface area and 210% increase per whole tissue. However, the affinity of the binding site for glucagon was not affected. The results suggested that an enhanced metabolic response of cold-acclimated rats to glucagon could be partly explained by the increased number of glucagon receptor in white adipocytes.
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PMID:Effect of cold acclimation on glucagon receptors of rat white adipocytes. 303 47

Guanine nucleotide and Mg2+ ion regulation of [125I-Tyr10]monoiodoglucagon ([125I]MIG) binding to liver plasma membranes from chicken, rat, and rabbit was studied. It was found that [125I]MIG binding to chicken liver membranes was increased by the addition of Mg2+ ion, while binding to rat and rabbit liver membranes was unaffected. In the chicken liver membranes, the Mg2+ ion induced high affinity binding which was sensitive to guanine nucleotides, while the low affinity binding in the absence of Mg2+ ion was not. Maximal effects of Mg2+ ion were observed at 1 mM. Glucagon binding to rat liver membrane receptors was GTP sensitive regardless of whether Mg2+ ion was added. Glucagon binding to rabbit liver membranes was insensitive to both Mg2+ ions and GTP. This lack of GTP effect was not due to degradation of GTP; no effect of the nonhydrolyzable analog guanyl-5'-yl-imidodiphosphate was observable. Glucagon stimulation of rabbit liver adenylyl cyclase, however, was dependent on GTP, as was the case with all of the other liver adenylyl cyclases studied here. The Kact of GTP for the rabbit liver system was very similar to that for rat liver membranes. The glucagon receptor was covalently labeled with [125I]MIG using p-hydroxysuccinimidyl azidobenzoate and analyzed by sodium dodecyl sulfate-gel electrophoresis. In all cases, a major labeled band at 63,000 daltons was observed. The levels of glucagon receptor and stimulatory (Ns) and inhibitory (Ni) regulatory proteins of adenylyl cyclase were measured. The highest levels of glucagon receptor were measured in rat liver membranes, while the levels in chicken and rabbit membranes were 30-40% lower. Rabbit liver membrane had the highest levels of Ns, while rat liver membranes had 2-fold lower and chick liver membrane 4-fold lower levels than rabbit liver membranes. The levels of Ni was similar in the three systems. Thus, the ratio of Ns to glucagon receptor was highest in the rabbit. In the rat, this ratio was 3-fold lower than that in the rabbit. In the chicken membranes, the ratio was about 60% of that in the rat. These data suggest that the observed differences in effects of GTP on hormone binding can be explained by alterations in the ratio of the receptor and Ns proteins among the various species.
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PMID:The hepatic glucagon receptor: a comparative study of the regulatory and structural properties. 303 85

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