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)

The hepatic glucagon receptor was covalently labeled with [125I-Try10]monoiodoglucagon [( 125I]MIG) by use of the heterobifunctional cross-linker hydroxysuccinimidyl p-azidobenzoate. Labeling of the Mr = 63,000 peptide was sensitive to glucagon and GTP at concentrations at which they affect [125I]MIG binding to the receptor. The labeled receptor was solubilized with Lubrol-PX, and the hydrodynamic characteristics of the receptor were determined. The molecular parameters of the solubilized receptor are: S20,w = 4.3 +/- 0.1, Stokes radius = 6.3 +/- 0.1 nm, frictional coefficient f/f0 = 1.8, and a calculated Mr = 119,000. Incubation of liver membranes at 32 degrees C for 15 min prior to the addition of [125I]MIG permitted us to identify the high molecular weight form (Mr = approximately 113,000) of the receptor by direct sodium dodecyl sulfate-gel electrophoretic analysis. The Mr = 63,000 peptide can be adsorbed to wheat germ lectin-Sepharose. The glycoprotein nature of the receptor has been utilized to develop an assay for the detergent-solubilized receptor that uses wheat germ lectin-Sepharose as a solid matrix to adsorb the [125I] MIG-receptor complex. The free hormone remains in the liquid phase and is removed in the supernatant after low speed centrifugation. 3-[(3-Cholamidopropyl)dimethylammonio]-1-propane sulfonate (CHAPS) solubilizes receptors with retention of [125I]MIG binding activity. [125I]MIG binding to the CHAPS-solubilized receptor is specifically affected by unlabeled glucagon. Interaction of [125I]MIG with the soluble receptor is insensitive to the presence of GTP. IC50 for glucagon using the soluble receptor was 33-70 nM, irrespective of the presence or absence of GTP, while when the membrane-bound receptor was used, the IC50 in the absence of GTP was 2-4 nM and in the presence of GTP was 35-80 nM. These data allow us to conclude that the hepatic glucagon receptor in the membrane and in the nondenaturing detergent solution is a dimer of the Mr = 63,000 hormone-binding subunit and a glycoprotein. The soluble receptor does not display any functional interaction with the stimulatory regulator.
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PMID:The hepatic glucagon receptor. Solubilization, characterization, and development of an affinity adsorption assay for the soluble receptor. 608 31

Insulin stimulates phosphorylation of both alpha- and beta- subunits of its own receptor in a cell-free system. A solubilized lectin-purified preparation of insulin receptors from rat liver membranes was preincubated with or without insulin at 4 degrees C and labeled for 10 min with Mn[gamma- 32P]ATP; the receptor subunits were isolated by specific immunoprecipitation with anti-receptor antibodies, followed by gel electrophoresis in sodium dodecyl sulfate. In gels run under reduced conditions, two bands (Mr = 135,000 and 95,000) were selectively labeled. These correspond exactly to the position of the alpha- and beta-subunits of the insulin receptor. Labeling of the Mr = 95,000 band was approximately 5-fold that of the Mr = 135,000 band. No labeled bands were detected when identical samples were immunoprecipitated in control serum. Phosphorylation of the receptor subunits required the presence of the divalent cation Mn2+ or Co2+; other cations such as Mg2+, Cr3+, Ca2+, and Zn2+ were ineffective. [gamma- 32P]ATP served as the 32P donor, whereas [gamma- 32P]GTP was ineffective. Phosphorylation of both subunits was stimulated 4-6-fold after a 60-min exposure to 10(-7) M pork insulin. Insulin-stimulated phosphorylation was half-maximal after 5 min of incubation with 10(-7) M insulin or after 18 h with 3 X 10(-10) M hormone. The enhanced phosphorylation was specific for insulin and its analogs; guinea pig insulin was about 2% as potent as pork insulin, whereas epidermal growth factor, adrenocorticotropic hormone, and glucagon, as well as cAMP, were ineffective. The rapidity and specificity of this reaction, as well as the presence of all necessary components in the plasma membrane, suggest that insulin-mediated receptor phosphorylation is one of the earliest biochemical steps following insulin binding.
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PMID:Characterization of insulin-mediated phosphorylation of the insulin receptor in a cell-free system. 633 57

High molecular weight glucagon immunoreactive material, obtained by gel-filtration (in the presence of 6 M guanidine hydrochloride) of fetal bovine pancreatic extracts, was tritiated by reductive methylation. Concanavalin-A-Sepharose column chromatography of the radiolabeled preparation yielded a discrete Concanavalin-A-reactive, alpha-methyl-mannoside-displaceable radioactive peak, coinciding with the glucagon immunoreactive peak. Submission of the Con-A-reactive material to wheat germ agglutinin-Sepharose column chromatography yielded a lectin-reactive, N-acetyl-glucosamine-displaceable radioactive peak, coinciding with the glucagon immunoreactive peak. The tritiated Con-A-reactive component interacted specifically with anti-glucagon antibodies. Sephacryl S-200 gel-filtration (in the presence of guanidine hydrochloride) dissociated a approximately 40 kDa radioactive species from the antibody-antigen complex. These data provide direct evidence for the existence of a large molecular weight glycosylated glucagon-related protein species from the fetal bovine pancreas.
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PMID:Characterization of a chemically tritiated large molecular weight glucagon immunoreactive protein species by lectin-affinity column chromatography and reaction with anti-glucagon antibodies. 654 29

Large glucagon immunoreactive substances, extracted from the fetal bovine pancreas and separated by gel filtration in the presence of 6 M guanidinium-hydrochloride, were submitted to lectin-sepharose affinity column chromatograph. Gel-filtered peak I (approximately 45 K delta) and peak II (approximately 10 K delta) interacted biospecifically with concanavalin-A- and wheat-germ-lectin-sepharoses, suggesting glycoproteins as possible constituents of large glucagon immunoreactive substances in extracts of the fetal bovine pancreas. The glucagon-like immunochemical identity of the lectin-sepharose-bound substances was further substantiated by binding to antiglucagon antibodies-sepharose and by characteristic proportional dilutions in the glucagon radioimmunoassay.
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PMID:Glycoprotein-like large glucagon immunoreactive species in extracts of the fetal bovine pancreas. 689 90

The glycoproteinic nature of the insulin receptor was indicated using two different approaches: 1. [125I] insulin binding to soluble receptors from mouse liver was inhibited by digestion with beta-galactosidase or pretreatment with Ricinus communis I or concanavalin A. An other enzyme (neuraminidase) and lectins (wheat germ agglutinin, Dolichos biflorus) did not affect the binding reaction. These data confirmed that insulin directly interacts with the galactoglycoproteins of liver membranes. 2. The galactose oxidase-sodium boro[3H] hydride technique, previously used for labeling accessible membrane galactoglycoproteins, was again utilized to discern the components that interact with insulin. When liver membranes were equilibrated with 10-7 M insulin prior to labeling, the SDS gel radioactive profiles were specifically modified with two galactoglycoprotein of apparent molecular sizes 195 000 and 145 000, compatible with their participation in the insulin binding interaction. Membrane pretreatment with beta-galactosidase or Sophora japonica lectin reduced the labeling in most peaks, thus supporting the argument for labeling sensitivity. Preincubation of membranes with 10-7 M proinsulin slightly hindered labeling, while pretreatment with 10-7 M glucagon was ineffective, suggesting a specificity of the insulin effect. These data indicate that glycoprotein nature of the insulin receptor for two reasons: alteration of insulin binding after modification of the galactoglycoproteins, and alteration of galactoglycoprotein labeling after insulin binding. Two galactoglycoproteins, with apparent molecular weights 145 000 and 195 000, respectively, were identified and they are suggested to have insulin binding properties.
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PMID:Identification of liver cell membrane galactoglycoproteins involved in the process of insulin binding. 703 Mar 99

The ability of pertussis toxin (PT) to recognize and bind to surface proteins on cells derived from pancreatic insulin-secreting beta cells and alpha cell-like glucagon-producing cells was investigated employing HIT-T15 (beta cell-derived) and In-R1-G9 (alpha cell-like) cell lines. PT recognition of membrane binding proteins on HIT-T15 and In-R1-G9 cells was first assessed with immunofluorescence microscopy in tissue culture. Both cell lines were equally well recognized by PT. N-octylglucoside extracts of whole cells and isolated membranes were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and blotted onto nitrocellulose membranes. PT, the B-oligomer, or the isolated PT dimers S2-S4 and S3-S4 recognized distinct proteins in HIT-T15 and In-R1-G9 cells of about 220 kDa. Recognition by the sialic acid specific Sambucus nigrica lectin identified these proteins as sialoglycoproteins. Incubation of the blotted membrane proteins with sialidase or pretreatment of PT with anti-PT polyclonal antibodies abolished the recognition and binding of these proteins by PT. To demonstrate that these glycoproteins are also able to transduce PT mediated effects and thus might serve as PT binding proteins, the stimulation of insulin secretion in HIT-T15 cells was assessed. As the secretion of insulin in HIT-T15 cells increased about 30% upon interaction with PT it was concluded that these glycoproteins are indeed functional as PT receptors.
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PMID:Identification of binding proteins for pertussis toxin on pancreatic beta cell-derived insulin-secreting cells. 756 12

Primary cultures of ovine ruminal epithelial cells were made to study the influence of collagen types I and IV out of medium supplementation with various hormones and Na-n-butyrate on cell morphology and growth characteristics. Both collagen type I and type IV led to increased cell proliferation with the stimulatory effect being more pronounced in collagen IV. In cultures grown on collagen I, both non-stratified and stratified colonies were found, whereas cultures grown on collagen IV showed predominantly stratified growth. Cells in both stratified and non-stratified colonies were positive for cytokeratin antibody. In non-stratified colonies, positive staining with fibronectin antibodies (FN-15) was found in a network over and around the cells. It is suggested that the non-stratified ruminal epithelial cells are in some respects similar to a 'non-differentiating keratinocyte' strain, derived from newborn foreskin epidermis. Cells in stratified colonies bound Ulex europaeus (UEAI) lectin which has been shown to be specific for differentiated epithelial cells in ruminal mucosa. Supplementation of culture medium with glucagon and insulin increased the total cell-overgrown area of collagen I cultures, whereas this effect was absent in cultures grown on collagen IV. In both cultures grown on collagen I or IV, hydrocortisone led to an increase in total cell-overgrown area, whereas Na-n-butyrate inhibited proliferation.
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PMID:Influences of extracellular matrix components on the growth and differentiation of ruminal epithelial cells in primary culture. 843 37

Neonatal porcine pancreases may be a potential source of islets for transplantation into patients with type 1 diabetes; however, whether these cellular grafts will be susceptible to damage by human natural antibody-mediated rejection remains controversial. Although we and others have demonstrated that porcine islets bind human IgG and IgM, it remains unknown if they express the xenoreactive antigen Gal alpha(1,3)Gal beta(1,4)GlcNAc-R (Gal epitope). In this study, by using the Gal-specific lectin IB4 for immunohistochemistry and fluorescence-activated cell sorter (FACS) analysis, we determined which cell types present in porcine neonatal islet cell (NIC) aggregates express the Gal epitope and which ones are susceptible to lysis by activation of the human complement. After FACS analysis, 30.0 +/- 3.0% of porcine NICs were shown to express Gal, whereas 70.0 +/- 2.0% did not. Histological assessment of Gal-expressing cells revealed that 54.9 +/- 8.8% stained positive for either insulin or glucagon. In contrast, 68.8 +/- 8.4% of the Gal-negative population stained positive for the pancreatic hormones insulin and glucagon. Incubation of either the Gal-positive or -negative cells with human AB serum plus complement for 1.5 h resulted in the lysis of >90% of the cells. These results demonstrate that porcine NIC aggregates are composed of Gal-expressing cells and that expression of Gal is not restricted to nonendocrine cells. Furthermore, both Gal-positive and Gal-negative cells are susceptible to human antibody/complement-mediated cytolysis, suggesting that this form of immunological destruction is an obstacle that will need to be overcome before porcine NIC aggregates can be used clinically.
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PMID:Expression of Gal alpha(1,3)gal on neonatal porcine islet beta-cells and susceptibility to human antibody/complement lysis. 972 28

Total parenteral nutrition (TPN) causes atrophy of gastrointestinal epithelia, so we asked whether lectins that stimulate epithelial proliferation can reverse this effect of TPN. Two lectins stimulate pancreatic proliferation by releasing CCK, so we asked whether lectins that stimulate gastrointestinal proliferation also release hormones that might mediate their effects. Six rats per group received continuous infusion of TPN and a once daily bolus dose of purified lectin (25 mg. rat-1. day-1) or vehicle alone (control group) for 4 days via an intragastric cannula. Proliferation rates were estimated by metaphase arrest, and hormones were measured by RIAs. Phytohemagglutinin (PHA) increased proliferation by 90% in the gastric fundus (P < 0.05), doubled proliferation in the small intestine (P < 0.001), and had a small effect in the midcolon (P < 0.05). Peanut agglutinin (PNA) had a minor trophic effect in the proximal small intestine (P < 0.05) and increased proliferation by 166% in the proximal colon (P < 0.001) and by 40% in the midcolon (P < 0.001). PNA elevated circulating gastrin and CCK by 97 (P < 0.05) and 81% (P < 0.01), respectively, and PHA elevated plasma enteroglucagon by 69% and CCK by 60% (both P < 0.05). Only wheat germ agglutinin increased the release of glucagon-like peptide-1 by 100% (P < 0.05). PHA and PNA consistently reverse the fall in gastrointestinal and pancreatic growth associated with TPN in rats. Both lectins stimulated the release of specific hormones that may have been responsible for the trophic effects. It is suggested that lectins could be used to prevent gastrointestinal atrophy during TPN. Their hormone-releasing effects might be involved.
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PMID:Gastrointestinal responses to a panel of lectins in rats maintained on total parenteral nutrition. 1033 15

The ATP-sensitive K(+) channel (K(ATP) channel) in pancreatic beta-cells is a critical regulator in insulin secretion. We previously reported that transgenic mice expressing a dominant-negative form (Kir6.2G132S) of Kir6.2, a subunit of the K(ATP) channel, specifically in beta-cells develop severe hyperglycemia in adults (8 weeks of age). In this study, we conducted a long-term investigation of the phenotype of these transgenic mice. Surprisingly, hyperglycemia was spontaneously improved with concomitant improvement of pancreatic insulin content in the transgenic mice at >25 weeks of age. Insulin-positive cells and pancreatic duodenal homeobox 1 (PDX1)-positive cells both were clearly increased in the older compared with the younger transgenic mice. Interestingly, cells labeled with the lectin Dolichos biflorus agglutinin (DBA), a potential indicator of uncommitted pancreatic epithelial/ductal cells, were detected in the islets of the transgenic mice but not in those of wild-type mice. In addition, a subset of the DBA-labeled cells was positive for PDX1, insulin, glucagon, somatostatin, or pancreatic polypeptide. Moreover, some of the DBA-labeled cells were also positive for a proliferating cell marker. These results show that the Kir6.2G132S transgenic mouse is a useful model for studying beta-cell regeneration and that DBA-labeled cells participate in the process.
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PMID:Spontaneous recovery from hyperglycemia by regeneration of pancreatic beta-cells in Kir6.2G132S transgenic mice. 1680 60


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