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At least 19 recordsLinked to original sources
PubMed1998

Dual glucagon recognition by pancreatic beta-cells via glucagon and glucagon-like peptide 1 receptors.

cAMP is required for normal glucose-induced insulin release by pancreatic beta-cells. In a previous study, we showed that cAMP production in beta-cells depends on the expression of receptors for glucagon, glucagon-like peptide 1(7-36) amide [GLP-1(7-36) amide], and glucose-dependent insulinotropic polypeptide. Although the latter two peptides are thought to amplify meal-induced insulin release (incretin effect), the role of glucagon in the regulation of insulin release remains elusive. In the present study, we analyzed the interaction of glucagon with its own receptor and with the glucagon-like peptide 1 (GLP-1) receptor using purified rat beta-cells. Glucagon binding was partially displaced by 1 micromol/l des-His1-[Glu9]glucagon-amide, a glucagon receptor antagonist, and by 1 micromol/l GLP-1. Conversely, GLP-1 binding was competitively inhibited by high glucagon concentrations (Ki = 0.3 micromol/l). Glucagon-induced cAMP production in beta-cells was inhibited both by 1 micromol/l des-His1-[Glu9]glucagon-amide and exendin-(9-39)-amide, a specific GLP-1 receptor antagonist, whereas GLP-1-induced cAMP formation was suppressed only by exendin-(9-39)-amide. Finally, addition of 1 micromol/l exendin-(9-39)-amide to 20 mmol/l glucose-stimulated beta-cells did not antagonize the potentiating effect of 1 nmol/l glucagon, although it prevented 45% of glucagon potentiation when the peptide was administered at 10 nmol/l. Our data suggest that glucagon recognition via two distinct receptors allows pancreatic beta-cells to detect this peptide both when diluted in the systemic circulation and when concentrated as local signal in the islet interstitium.

Animals↗
PubMed1980

Preparation of 2-thioltryptophan-glucagon and (tryptophan-S-glucagon)2. Differences in binding to the glucagon receptor in the hepatic adenylate cyclase system.

The synthesis of 2-thioltryptophan-glucagon is described. Oxidation of this compound gives the dimer (Trp-S-glucagon)2. Both monomer and dimer are equi-potent on a molar basis with native glucagon as activators of adenylate cyclase in hepatic plasma membranes. However, from the ability to compete with 125I-glucagon for binding at the glucagon receptor, the dimer has one-fourth the binding affinity for the receptor as does native glucagon, 2-thiol-Trp-glucagon, and Trp-(2,4-dinitrophenylsulfenyl)-glucagon which have equal affinities for the receptor. Addition of GTP, which converts the receptor from a tight-binding to a lower affinity form and which activates adenylate cyclase in the presence of glucagon, allows (Trp-S-glucagon)2 to bind equally with native glucagon and the other thiol derivatives of the hormone. This effect of GTP on the binding of the glucagon dimer and the uses of 2-thiol-Trp-glucagon in the semisynthesis of new glucagon derivatives are discussed.

Adenylyl Cyclases↗
PubMed2002

The role of salt bridge formation in glucagon: an experimental and theoretical study of glucagon analogs and peptide fragments of glucagon.

BACKGROUND: Glucagon is a 29-residue peptide produced in the alpha cells of the pancreas that interacts with hepatic receptors to stimulate glucose production and release, via a cAMP-mediated pathway. Type 2 diabetes patients may have an excess of glucagon and, as such, glucagon antagonists might serve as diabetes drugs. The antagonists that bind to the glucagon receptor but do not exhibit activity could be analogs of glucagon. The presence of salt bridges between some residues of glucagons (such as aspartic acid) and others (such as lysine) might influence both the binding to the receptor and the activity. MATERIALS AND METHODS: Experimental-The solid phase method with 4-methylbenzilhydrilamine resin (p-MBHA resin) was used for the synthesis of glucagon analogs. Rat liver membranes were prepared from male Sprague-Dawley rats by the Neville procedure. The receptor binding essay was performed in 1% BSA, 1 mM dithiothreitol, 25 mM Tris-HCl buffer, pH 7.2. Adenyl cyclase activity was measured in an assay medium containing 1% serum albumin, 25 mM MgCl2, 2 mM dithiothreitol, 0.025 mM GTP, 5 mM ATP, 0.9 mM theophylline, 17.2 mM creatine phosphate, and 1 mg/ml creatine phosphokinase. Theoretical-Quantum chemical calculations using the Titan program with the 6-31G* basis set were performed to calculate the binding energies of salt bridges between aspartic or glutamic acids and lysine. The relative stability of cyclic conformations of glucagon segments versus the extended segments was determined. RESULTS: It was found that the cyclic Glu9-Lys12 amide compound displayed a 20-fold decrease in binding affinity. DesHis1 cyclic compounds Glu20-Lys24 amide and DesHis1Glu9 Glu20-Lys24 amide behave as glucagon antagonists. The calculations show that cyclic conformations of tetrapeptidic and pentapeptidic segments of glucagon are more stable than the extended species. CONCLUSIONS: The biological data and the theoretical calculations show that an intramolecular salt bridge might impart stability to some glucagon antagonists and, when situated at the C-terminus of glucagon, might facilitate induction of an alpha-helix upon initial hormone association with the membrane bilayer. These findings might be a useful tool for the design of new glucagon antagonists.

Animals↗
PubMed1981

Chromatographic pattern of extrapancreatic glucagon and glucagon-like immunoreactivity before and during stimulation by epinephrine and participation of glucagon in epinephrine-induced hepatic glucose overproduction.

To characterize the glucagon released in response to epineephrine in depancreatized dogs, plasma samples before and during epinephrine infusion were subjected to molecular-sieve chromatography on Bio-Gel P-30 columns. The chromatographic profile for extrapancreatic immunoreactive glucagon (eIRG) revealed two glucagon moieties of molecular weight 9,000 to 12,000. GLI of this molecular weight was released in response to epinephrine only under conditions of prevailing hyperglycemia. To determine if glucagon's participation in epinephrine-induced hepatic glucose overproduction in diabetes was dependent upon the degree of metabolic control, six conscious depancreatized dogs were infused with epinephrine or epinphrine plus somatostatin, under conditions of prevailing hyperglycemia or normoglycemia. Under normoglycemic conditions, epinephrine stimulated eIRG release, but there was a similar rise in hepatic glucose production (Ra) with or without glucagon suppression by somatostatin. Under hyperglycemic conditions, epinephrine stimulated eIRG and GLI release, and the rise in Ra was significantly greater with epinephrine than with epinephrine plus somatostatin infusion. Thus, under conditions of good metabolic control, epinephrine increased hepatic glucose production independently of glucagon, whereas with poor metabolic control, glucagon contributed to hepatic overproduction of glucose.

Animals↗
PubMed1988

Isolation of alligator gar (Lepisosteus spatula) glucagon, oxyntomodulin, and glucagon-like peptide: amino acid sequences of oxyntomodulin and glucagon-like peptide.

Oxyntomodulin, glucagon, and a glucagon-like peptide (GLP) have been isolated from the endocrine pancreas of the alligator gar (Lepisosteus spatula), a ganoid fish. The three peptides were isolated by gel filtration and HPLC and were identified by size, composition, and glucagon-like immunoreactivity. The amino acid sequences of the oxyntomodulin and GLP were determined. The oxyntomodulin contains 36 amino acid residues and its sequence is H S Q G T F T N D Y S K Y L D T R R A Q D F V Q W L M S T K R S G G I T. The composition of the glucagon is identical to the N-terminal 29 residues of the gar oxyntomodulin. The single form of GLP found contains 34 amino acid residues in the following sequence: H A D G T Y T S D V S S Y L Q D Q A A K K F V T W L K Q G Q D R R E. These findings suggest that all three peptides are derived from a common precursor.

Amino Acid Sequence↗
PubMedDate not supplied

Glucagon-37 (oxyntomodulin) and glucagon-29 (pancreatic glucagon) in human bowel: analysis by HPLC and radioreceptorassay.

A method for assaying specifically the biologically active peptides Glucagon-37 (G-37/Oxyntomodulin/bioactive Enteroglucagon) and Glucagon-29 (G-29/pancreatic Glucagon) has been developed by use of high performance liquid chromatography (HPLC) of crude tissue extracts followed by radioreceptorassay in liver membranes. The peaks observed with this method in samples from human bowel have also been analysed in two other assays: stimulation of cyclic AMP accumulation in gastric glands and radioimmunoassay. Owing to the different patterns of activity of porcine G-37 and G-29 in these assays, the comparison of the data obtained allows to discriminate between the two peptides. The same behaviour in both HPLC and the three assays of the human peaks on one hand and the porcine peptides on the other strongly suggests that human intestine contains a very similar or the same molecules as that isolated from the porcine tissues. Whatever the portion of small intestine, G-37 represented ca 90% of G-37 + G-29. A decreasing concentration gradient of both G-37 and G-29 was also observed from ileum to descending colon.

Animals↗
PubMed2000

Dipeptidyl peptidase IV (DPIV/CD26) degradation of glucagon. Characterization of glucagon degradation products and DPIV-resistant analogs.

Over the past decade, numerous studies have been targeted at defining structure-activity relationships of glucagon. Recently, we have found that glucagon(1-29) is hydrolyzed by dipeptidyl peptidase IV (DPIV) to produce glucagon(3-29) and glucagon(5-29); in human serum, [pyroglutamyl (pGlu)(3)]glucagon(3-29) is formed from glucagon(3-29), and this prevents further hydrolysis of glucagon by DPIV (H.-U. Demuth, K. Glund, U. Heiser, J. Pospisilik, S. Hinke, T. Hoffmann, F. Rosche, D. Schlenzig, M. Wermann, C. McIntosh, and R. Pederson, manuscript in preparation). In the current study, the biological activity of these peptides was examined in vitro. The amino-terminally truncated peptides all behaved as partial agonists in cyclic AMP stimulation assays, with Chinese hamster ovary K1 cells overexpressing the human glucagon receptor (potency: glucagon(1-29) > [pGlu(3)]glu- cagon(3-29) > glucagon(3-29) > glucagon(5-29) > [Glu(9)]glu- cagon(2-29)). In competition binding experiments, [pGlu(3)]glucagon(3-29) and glucagon(5-29) both demonstrated 5-fold lower affinity for the receptor than glucagon(1-29), whereas glucagon(3-29) exhibited 18-fold lower affinity. Of the peptides tested, only glucagon(5-29) showed antagonist activity, and this was weak compared with the classical glucagon antagonist, [Glu(9)]glucagon(2-29). Hence, DPIV hydrolysis of glucagon yields low affinity agonists of the glucagon receptor. As a corollary to evidence indicating that DPIV degrades glucagon (Demuth, et al., manuscript in preparation), DPIV-resistant analogs were synthesized. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry was used to assess DPIV resistance, and it allowed kinetic analysis of degradation. Of several analogs generated, only [D-Ser(2)] and [Gly(2)]glucagon retained high affinity binding and biological potency, similar to native glucagon in vitro. [D-Ser(2)]Glucagon exhibited enhanced hyperglycemic activity in a bioassay, whereas [Gly(2)]glucagon was not completely resistant to DPIV degradation.

Animals↗
PubMed1982

Structure-conformation-activity studies of glucagon and semi-synthetic glucagon analogs.

Examination of glucagon structure-activity relationships and their use for the development of glucagon antagonists (inhibitors) have been hampered until recently by the lack of high purity of semisynthetic glucagon analogs and inadequate study of full dose-response curves for these analogs in sensitive bioassay systems. Recently a number of highly purified glucagon fragments and semi-synthetic analogs have been prepared and their full dose-response activities examined over a wide concentration range using the hepatic membrane adenylate cyclase assay, the hepatic membrane receptor binding assay, and glycogenolytic activity in isolated rat hepatocytes. The results of these studies have enabled us to identify and dissociate the structural (and in some cases conformational) features of glucagon important for binding from those most responsible for biological activity (transduction). Key findings in these studies were the observation that: (1) the C-terminal region of glucagon is primarily of importance for hormone binding to receptors; (2) glucagon 1-21 and glucagon 1-6 have low potency, but are essentially fully active glucagon derivatives; and (3) highly purified glucagon 2-29 ([1-des-histidine]-glucagon), [1-N alpha-carbamoylhistidine]-glucagon and [1-N alpha-carbamoylhistidine, 12-N alpha-carbamoyllysine]-glucagon are all partial agonists. These and other findings led us to synthesize several semisynthetic analogs of glucagon which were found to possess no intrinsic biological activity in the hepatic adenylate cyclase assay system, but which could block the effect of glucagon (competitive inhibitors) in activating adenylate cyclase in this system. Two of these highly purified analogs [1-des-histidine][2-N alpha-trinitrophenylserine, 12-homoarginine]-glucagon and [1-N alpha-trinitrophenylhistidine, 12-homoarginine]-glucagon were quite potent glucagon antagonists (inhibitors) with pA2 values of 7.41 and 8.16 respectively. The latter compound has also been demonstrated to decrease dramatically blood glucose levels of diabetic animals in vivo. These results demonstrate that glucagon is a major contributor to the hyperglycemia of diabetic animals. Examination of the known and calculated conformational properties of glucagon provide insight into the structural and conformational properties of glucagon and its analogs most responsible for its biological activity. Consideration of these features and the mechanism of glucagon action at the membrane receptor level provide a framework for further developing glucagon analogs for theoretical and therapeutic applications.

Adenylyl Cyclases↗
PubMed1982

Inhibition of glucagon secretion by exogenous glucagon in the isolated, perfused dog pancreas.

To determine if glucagon can inhibit its own secretion, exogenous glucagon was perfused in the isolated canine pancreas at concentrations ranging from 90 to 1050 pg/ml during either stimulation or suppression of endogenous glucagon secretion. When endogenous glucagon secretion was completely suppressed by the addition of 200 mg/dl of glucose to the perfusate, the concentration of glucagon in the venous effluent ranged from 80.8 to 89.9% of the level perfused; these deficits of 10-20% were attributed to uptake and/or degradation of exogenous glucagon by the pancreas. By contrast, when endogenous glucagon secretion was stimulated by perfusing with 10 mM arginine or by glucopenia of 25 mg/dl, there was a significant increase in the glucagon deficit (the efflux glucagon level during glucagon perfusion minus the sum of the pre-perfusion efflux level of endogenous glucagon and the concentration of glucagon perfused); only 60-76% of the expected glucagon concentration was present in the venous efflux. This increase in deficit is assumed to reflect suppression of endogenous glucagon. The glucagon deficit rose progressively with deficit reached the approximate level of endogenous glucagon, at which no further increase occurred, suggesting that suppression at that point was complete. To determine if the glucagon-suppressing action of glucagon was mediated by an increase in glucagon-stimulated insulin or somatostatin secretion, these hormones were measured in the various experiments. Insulin and somatostatin increased significantly when glucagon was perfused with the glucose arginine-containing solution but did not rise significantly when the glucopenic buffer was used, although suppression of endogenous glucagon was similar in each instance. It is concluded that glucagon suppresses glucagon secretion. Evidence that insulin and/or somatostatin mediate this action could not be obtained.

Animals↗
PubMed1990

Glucagon-(19-29), a Ca2+ pump inhibitory peptide, is processed from glucagon in the rat liver plasma membrane by a thiol endopeptidase.

Glucagon-(19-29) is 1000-fold more potent that glucagon as an inhibitor of the liver plasma membrane calcium pump, which suggests that this peptide fragment is naturally occurring. Since glucagon-(19-29) is undetectable in plasma, the processing of glucagon into its (19-29) fragment may occur upon interaction of glucagon with its target tissues. The use of a specific radioimmunoassay for glucagon-(19-29) in association with the separation and identification of peptides by high performance liquid chromatography revealed that, upon incubation at 37 degrees C with hepatic plasma membranes, glucagon is processed into its (19-29) C-terminal fragment. The identity of the fragment was confirmed by amino acid sequencing. The processing activity was inhibited by reagents of the thiol group and by 1,10-phenanthroline, suggesting that a thiol endopeptidase containing a catalytically active metal is involved in this processing. Following its production, glucagon-(19-29) was degraded with a half-life of less than 10 s. This degradation was inhibited by bacitracin and by the aminopeptidase inhibitors bestatin and amastatin. When glucagon was incubated with liver plasma membranes in the absence of inhibitors, the accumulation of glucagon-(19-29) reached a maximum at 2 min (1% of initial glucagon), followed by a slow decline. In the presence of bacitracin and bestatin, the amounts of glucagon-(19-29) obtained from glucagon increased continuously, 1 and 2% of glucagon being transformed after 10 and 30 min, respectively. The production of glucagon-(19-29) did not appear to be associated with the binding of glucagon to its receptors, since (i) guanosine 5'-(3-O-thio)triphosphate, a compound which decreases the glucagon-receptor interaction, could not decrease the conversion of glucagon into glucagon-(19-29); (ii) a glucagon analogue which displays a strongly decreased affinity for the hepatic glucagon receptors was processed similarly to glucagon. The conversion also occurs upon incubation with intact hepatoma cells in monolayer culture. These observations suggest that, under physiological conditions, glucagon is processed in liver by cleavage of the Arg17-Arg18 basic doublet, leading to the production of a fragment which is known to display an original biological specificity, namely the modulation of the hepatocyte plasma membrane calcium pump.

Adenosine Triphosphate↗
PubMed2004

Effects of positive and negative lens treatment on retinal and choroidal glucagon and glucagon receptor mRNA levels in the chicken.

PURPOSE: It has been found in the chicken that the amount of retinal glucagon mRNA increases during treatment with positive lenses. Pharmacological studies support the idea that glucagon may act as a stop signal for visually induced eye growth. To gain more insight into the functional role of glucagon, the changes of glucagon and glucagon receptor mRNA concentrations in retina and choroid over time were studied. Furthermore, the abundance of glucagon and the glucagon receptor was studied in different fundal layers (retina, retinal pigment epithelium[RPE], choroid) and the blood. METHODS: Semiquantitative real-time RT-PCR was used to measure glucagon and glucagon receptor mRNA levels in retina and choroid after positive and negative lens treatment for 2, 6, or 24 hours, by unilateral -7- or +7-D lenses. Contralateral eyes served as the control, and completely untreated animals provided further reference data. Intravitreal colchicine injections (which are known to reduce the number of glucagon cells sharply) were used to verify that the related decline in glucagon mRNA could be measured by real-time RT-PCR. RESULTS: In the retina, treatment with -7-D lenses induced an initial upregulation of glucagon mRNA in both eyes, followed by a significant downregulation. The treatment with +7-D lenses showed a significant but transient downregulation in the control eye superimposed on a trend toward upregulation in the treated eye. However, the changes in glucagon mRNA expression were not confined to the lens-treated eyes but were also found, although sometimes to a lesser extent, in the non-lens-covered fellow eyes. There was evidence of a transient increase in glucagon receptor mRNA levels in lens-treated eyes after either -7- or +7-D lens wear. In the choroid, no effect of imposed defocus was detected. The injection of colchicine led to the destruction of approximately 75% of the glucagon amacrine cells but the mRNA level of retinal glucagon decreased by only approximately 50%. Glucagon receptor expression was found to be higher in the RPE than the retina and choroid whereas, in the blood, glucagon and glucagon receptor mRNA expression was below detection level. CONCLUSIONS: The observed bidirectional regulation of glucagon mRNA in correlation with the sign of imposed defocus supports the idea that glucagon may act as a stop-and-go signal for eye growth. This is in line with a previous proposal based on studies of changes of the glucagon peptide content.

Animals↗
PubMed1971

The reaction of glucagon with its receptor: evidence for discrete regions of activity and binding in the glucagon molecule.

Des-histidine-glucagon (DH-glucagon, glucagon(2-29)) does not activate the glucagon-sensitive adenylate cyclase system present in either liver plasma membranes or in fat-cell "ghosts", but inhibits the response of these systems to submaximal concentrations of glucagon. DH-glucagon also inhibits, competitively, the binding of [(125)I]glucagon to its receptor in liver plasma membranes. Amino-terminal fragments of glucagon (glucagon(1-21), glucagon(1-23)) and carboxy-terminal fragments (glucagon(20-29), glucagon(22-29)) failed to activate adenylate cyclase, to inhibit the response of the enzyme to glucagon, or to compete with labeled glucagon at its receptor. It is concluded that the amino-terminal histidine residue of glucagon is essential for biological activity and that a hydrophobic near-carboxy-terminal region (residues 22-27) is essential for binding of glucagon to its receptor. Amino-terminal histidine may also contribute to the binding of glucagon, since the apparent affinity of DH-glucagon for the receptor is only about one-sixth that of glucagon. Thus, essentially the entire molecule of glucagon must be considered to be the biologically active species.Because, as shown elsewhere, the binding of glucagon to its receptor shows characteristics of hydrophobic bonding, and because certain detergents induce conformational changes in the carboxy-terminal binding region of glucagon, the binding is probably of a lipophilic type.

Adenylyl Cyclases↗
PubMed2004

Hepatic glucagon receptor binding and glucose-lowering in vivo by peptidyl and non-peptidyl glucagon receptor antagonists.

Glucagon receptor antagonists have been actively pursued as potential therapeutics for the treatment of type 2 diabetes. Peptidyl and non-peptidyl glucagon receptor antagonists have been shown to block glucagon-induced blood glucose elevation in both animals and humans. How the antagonists and the glucagon receptor interact in vivo has not been reported and is the subject of the current study. Using (125)I-labeled glucagon as a radiotracer, we developed an in vivo glucagon receptor occupancy assay in mice expressing a human glucagon receptor in place of the endogenous mouse glucagon receptor (hGCGR mice). Using this assay, we first showed that the glucagon receptor is expressed predominantly in liver, to a much lesser extent in kidney, and is below detection in several other tissues/organs in the mice. We subsequently showed that, at 2 mg/kg body weight (mg/pk) dosed intraperitoneally (i.p.), peptidyl glucagon receptor antagonist des-His-glucagon binds to approximately 78% of the hepatic glucagon receptor and blocks an exogenous glucagon-induced blood glucose elevation in the mice. Finally, we also showed that, at 10 and 30 mg/kg dosed orally (p.o.), compound A, a non-peptidyl small molecule glucagon receptor antagonist, occupied 65-70% of the hepatic glucagon receptor, and significantly diminished exogenous glucagon-induced blood glucose elevation in the mice. At 3 mg/kg, however, compound A occupied only approximately 39% of the hepatic glucagon receptor and did not affect exogenous glucagon-induced blood glucose elevation in the mice. Taken together, the results confirmed previous reports that glucagon receptors are present predominantly in the liver, and provide the first direct evidence that peptidyl and non-peptidyl glucagon receptor antagonists bind to the hepatic glucagon receptor in vivo, and that at least 60% receptor occupancy correlates with the glucose lowering efficacy by the antagonists in vivo.

Animals↗
PubMed1997

The role of phenylalanine at position 6 in glucagon's mechanism of biological action: multiple replacement analogues of glucagon.

Extensive evidence gathered from structure-activity relationship analysis has identified and confirmed specific positions in the glucagon sequence that are important either for binding to its receptor or for signal transduction. Fifteen glucagon analogues have been designed and synthesized by incorporating structural changes in the N-terminal region of glucagon, in particular histidine-1, phenylalanine-6, and aspartic acid-9. This investigation was conducted to study the role of phenylalanine at position 6 on the glucagon mechanism of action. These glucagon analogues have been made by either deleting or substituting hydrophobic groups, hydrophilic groups, aromatic amino acids, or a D-phenylalanine residue at this position. The structures of the new analogues are as follows: [des-His1, des-Phe6, Glu9]glucagon-NH2 (1); [des-His1,Ala6,Glu9]glucagon-NH2 (2); [des-His1,Tyr6,Glu9]glucagon-NH2 (3); [des-His1,Trp6,Glu9]-glucagon-NH2 (4); [des-His1,D-Phe6,Glu9]glucagon-NH2 (5); [des-His1,Nle6,Glu9]glucagon-NH2 (6); [des-His1,Asp6,Glu9]glucagon-NH2 (7); [des-His1,des-Gly4,Glu9]glucagon-NH2 (8); [desPhe6,-Glu9]glucagon-NH2 (9); [des-Phe6]glucagon-NH2 (10); [des-His1, des-Phe6]glucagon-NH2 (11); [des-His1, des-Phe6,Glu9]glucagon (12); [des-Phe6,Glu9]glucagon (13); [des-Phe6]glucagon (14); and [des-His1, des-Phe6]glucagon (15). The receptor binding potencies IC50 values are 48 (1), 126 (2), 40 (3), 19 (4), 100 (5), 48 (6), 2000 (7), 52 (8), 113 (9), 512 (10), 128 (11), 1000 (12), 2000 (13), 500 (14), and 200 nM (15). All analogues were found to be antagonists unable to activate the adenylate cyclase system even at concentrations as high as 10(-5) M except for analogues 6 and 8, which were found to be weak partial agonists/partial antagonists with maximum stimulation between 6-12%. In competitive inhibition experiments, all the analogues caused a right shift of the glucagon-stimulated adenylate cyclase dose-response curve. The pA2 values were 8.20 (1), 6.40 (2), 6.20 (3), 6.25 (4), 6.30 (5), 6.30 (7), 6.05 (8), 6.20 (9), 6.30 (10), 6.25 (11), 6.10 (12), 6.20 (13), 6.20 (14), and 6.35 (15).

Adenylyl Cyclases↗
PubMed2003

Three distinct epitopes on the extracellular face of the glucagon receptor determine specificity for the glucagon amino terminus.

The glucagon and glucagon-like peptide-1 (GLP-1) receptors are homologous family B seven-transmembrane (7TM) G protein-coupled receptors, and they selectively recognize the homologous peptide hormones glucagon (29 amino acids) and GLP-1 (30-31 amino acids), respectively. The amino-terminal extracellular domain of the glucagon and GLP-1 receptors (140-150 amino acids) determines specificity for the carboxyl terminus of glucagon and GLP-1, respectively. In addition, the glucagon receptor core domain (7TM helices and connecting loops) strongly determines specificity for the glucagon amino terminus. Only 4 of 15 residues are divergent in the glucagon and GLP-1 amino termini; Ser2, Gln3, Tyr10, and Lys12 in glucagon and the corresponding Ala8, Glu9, Val16, and Ser18 in GLP-1. In this study, individual substitution of these four residues of glucagon with the corresponding residues of GLP-1 decreased the affinity and potency at the glucagon receptor relative to glucagon. Substitution of distinct segments of the glucagon receptor core domain with the corresponding segments of the GLP-1 receptor rescued the affinity and potency of specific glucagon analogs. Site-directed mutagenesis identified the Asp385 --> Glu glucagon receptor mutant that specifically rescued Ala2-glucagon. The results show that three distinct epitopes of the glucagon receptor core domain determine specificity for the N terminus of glucagon. We suggest a glucagon receptor binding model in which the extracellular ends of TM2 and TM7 are close to and determine specificity for Gln3 and Ser2 of glucagon, respectively. Furthermore, the second extracellular loop and/or proximal segments of TM4 and/or TM5 are close to and determine specificity for Lys12 of glucagon.

Amino Acid Sequence↗