scholarly journals Guanosine nucleotides regulate hormone binding of insulin receptors

1992 ◽  
Vol 281 (3) ◽  
pp. 735-743 ◽  
Author(s):  
E R Mortensen ◽  
J Drachman ◽  
G Guidotti
Keyword(s):  
Binding Sites ◽  
Plasma Membranes ◽  
Insulin Receptors ◽  
Insulin Binding ◽  
Scatchard Analysis ◽  
Hormone Binding ◽  
Temperature Dependent ◽  
High Affinity ◽  
Mild Reduction ◽  
Adipocyte Plasma Membranes

Insulin receptors in turkey erythrocyte and rat adipocyte plasma membranes display non-linear hormone binding by Scatchard analysis. This result is consistent with evidence that the insulin-binding sites are heterogeneous and have at least two affinities for the hormone. Mild reduction of plasma membranes with dithiothreitol, before insulin binding, increased the fraction of hormone binding with high affinity without significantly changing the total number of receptor-binding sites. In the presence of guanosine 5′-[gamma-thio]triphosphate, the amount of receptor with high affinity for insulin in the reduced membranes decreased to that present in the absence of reduction; the effect of the nucleotide was concentration- and temperature-dependent. This decrease in insulin binding was specific for guanine nucleotides.

Desensitization of beta-adrenergic receptors in adipocytes causes increased insulin sensitivity of glucose transport

1996 ◽  
Vol 271 (2) ◽  
pp. E271-E276 ◽  
Author(s):  
A. Green ◽  
R. M. Carroll ◽  
S. B. Dobias
Keyword(s):  
Insulin Sensitivity ◽  
Glucose Transport ◽  
Adrenergic Receptors ◽  
Plasma Membranes ◽  
Insulin Receptors ◽  
Insulin Binding ◽  
High Dose ◽  
Beta Adrenergic Receptors ◽  
Beta Adrenergic ◽  
Adipocyte Plasma Membranes

To determine the effect of desensitization of adipocyte beta-adrenergic receptors on insulin sensitivity, rats were continuously infused with isoproterenol (50 or 100 micrograms.kg-1.h-1) for 3 days by osmotic minipumps. Epididymal adipocytes were isolated. The cells from treated animals were desensitized to isoproterenol, as determined by response of lipolysis (glycerol release). Binding of [125I]iodocyanopindolol was decreased by approximately 80% in adipocyte plasma membranes isolated from treated rats, indicating that beta-adrenergic receptors were downregulated. Cellular concentrations of Gn alpha and Gi alpha were not altered. Insulin sensitivity was determined by measuring the effect of insulin on glucose transport (2-deoxy-[3H]glucose uptake). Cells from the isoproterenol-infused rats were markedly more sensitive to insulin than those from control rats. This was evidenced by an approximately 50% increase in maximal glucose transport rate in cells from the high-dose isoproterenol-treated rats and by an approximately 40% decrease in the half-maximal effective concentration of insulin in both groups. 125I-labeled insulin binding to adipocytes was not altered by the isoproterenol infusions, indicating that desensitization of beta-adrenergic receptors results in tighter coupling between insulin receptors and stimulation of glucose transport.

Chronic and endogenous regulation of insulin receptors by catecholamines in adipocytes from patients with a phaeochromocytoma

1990 ◽  
Vol 10 (2) ◽  
pp. 201-207 ◽  
Author(s):  
M. C. Carranza ◽  
M. A. Simón ◽  
A. Torres ◽  
C. Calle
Keyword(s):  
Adipose Tissue ◽  
Binding Sites ◽  
Insulin Receptors ◽  
Human Adipose Tissue ◽  
Insulin Binding ◽  
Scatchard Analysis ◽  
Cooperative Model ◽  
Receptor Affinity ◽  
Site Model ◽  
Binding Data

Insulin binding in adipocytes from patients with a phaeochromocytoma (PH) approached that of the controls (C) at low and higher concentrations of unlabeled insulin. The apparent receptor affinity was unchanged (ED50: PH 0.50×10−9M and C0.60×10−9M). Scatchard analysis of the binding data using the negative cooperative model revealed a 46% decrease in the total number of receptors together with no changes in both K−e (PH 0.55×109M−1 and C 0.36×109M−1) and K−f (PH 0.13×109 M−1 and C 0.07×109 M−1). According to the two site model, an altered proportion in the two classes of insulin binding sites was detected. This was accompanied by a catecholamine-desensitization of the adipocytes to the antilipolytic action of insulin. These events could represent a final situation of a chronic and endogeneous regulation by high levels of catecholamines of insulin receptors in human adipose tissue.

Adipocyte insulin-binding species: the 40 Å Stoke's radius protein

1984 ◽  
Vol 62 (7) ◽  
pp. 566-570
Author(s):  
C. Elliott ◽  
H. Joseph Goren
Keyword(s):  
Insulin Receptor ◽  
Gel Electrophoresis ◽  
Plasma Membranes ◽  
Insulin Receptors ◽  
Insulin Binding ◽  
Relative Mass ◽  
Single Polypeptide Chain ◽  
Dodecyl Sulfate ◽  
Single Polypeptide ◽  
Adipocyte Plasma Membranes

Several laboratories have demonstrated the presence of large (70 Å) (1 Å = 0.1 nm) and small (40 Å) insulin receptors. This report provides evidence that the 40 Å insulin receptor migrates on dodecyl sulfate – acrylamide gel electrophoresis as a 90 000 dalton protein and that this protein is a single polypeptide chain. 125I-labeled insulin was bound to plasma membranes from isolated rat adipocytes. Following removal of unbound 125I-labeled insulin, the mixture was exposed to disuccinimidyl suberate. Proteins tagged with 125I-labeled insulin were separated by dodecyl sulfate gel electrophoresis or Sepharose 6B chromatography. Autoradiography of the gels demonstrated several large (relative mass (Mr) > 300 000) and one small (Mr ~ 90 000) labeled protein in nonreduced membrane proteins. Dithiothreitol reduction decreased the large insulin-binding species to its known subunits, but the 90 000 dalton protein did not decrease in size. Triton X-100 solubilized plasma membranes were separated by Sepharose 6B chromatography. One labeled protein, with Kav = 0.57 elution position, on dodecyl sulfate gel electrophoresis migrated as a 90 000 dalton protein. Thus, rat adipocyte plasma membranes contain both an oligomeric insulin-binding species and a monomeric insulin-binding species. The relationship of the monomeric to the oligomeric insulin receptor is discussed.

THE BINDING OF INSULIN AND SOMATOMEDIN A TO HUMAN PLACENTAL MEMBRANE

1975 ◽  
Vol 80 (1) ◽  
pp. 14-31 ◽  
Author(s):  
K. Takano ◽  
K. Hall ◽  
L. Fryklund ◽  
A. Holmgren ◽  
H. Sievertsson ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Binding Sites ◽  
Membrane Fraction ◽  
Specific Binding ◽  
Insulin Binding ◽  
Radioreceptor Assay ◽  
Affinity Constant ◽  
Scatchard Analysis ◽  
Mean Values ◽  
High Affinity

ABSTRACT A particulate membrane fraction from human placental membrane was shown to be rich in binding sites not only for insulin but also for somatomedin A. The binding of the 125I-labelled peptide was time and temperature dependent. Degrading activity present in the membrane fraction was negligible at +4°C. The Scatchard plot for insulin binding revealed two types of binding sites with an apparent high affinity constant of 3.8× 108 m−1 and with 5.4 × 10−9 moles of binding sites per mg of membrane protein. The Scatchard analysis of somatomedin A revealed two classes of binding sites with an apparent high affinity constant of 2.7 × 107 m−1 and with 1.9× 10−8 moles of binding sites per mg of membrane protein. In high concentrations insulin interfered with the specific binding sites for somatomedin A and vice versa. In comparison with insulin the somatomedin A preparation was one million times more potent in displacing labelled somatomedin A than in displacing labelled insulin from their respective binding sites. A radioreceptor assay utilizing particulate placental membrane and labelled somatomedin A purified on the membrane enabled the determination of somatomedin in unextracted serum. The mean values of somatomedin A in sera from patients with pituitary dwarfism and acromegaly were 0.57 and 3.2 U/ml, respectively by radioreceptor assay and 0.41 and 1.61 U/ml, respectively by bioassay. Various causes of this discrepancy between the methods are discussed.

PGE2 binding, synthesis, and distribution in hen oviduct

1985 ◽  
Vol 248 (1) ◽  
pp. E80-E88
Author(s):  
G. Asboth ◽  
H. Todd ◽  
M. Toth ◽  
F. Hertelendy
Keyword(s):  
Binding Sites ◽  
Scatchard Analysis ◽  
Affinity Binding ◽  
Shell Gland ◽  
Temperature Dependent ◽  
Single Class ◽  
High Affinity Binding ◽  
High Affinity ◽  
Prostaglandin F2 Alpha ◽  
20 Nm

Prostaglandin E2 (PGE2) bound specifically to particulate fractions prepared from the vagina and uterus (shell gland) portions of the hen oviduct in a time and temperature dependent fashion. Scatchard analysis indicated a single class of high-affinity binding sites in the vagina (Kd congruent to 1 nM), whereas the myometrium exhibited two kinds of binding site populations (Kd1 congruent to 1 nM, Kd2 congruent to 20 nM). It is suggested that these binding sites represent specific PGE2 receptors mediating the effects of PGE2 in oviductal smooth muscle. Vaginal particulate fractions produced approximately four times more prostanoids from [3H]-arachidonate than did uterine preparations. In the presence of epinephrine both tissues synthesized mainly thromboxane (TxB2), PGE2, and significantly less prostaglandin F2 alpha (PGF2 alpha). Addition of glutathione (GSH) or cytosol prepared from the oviduct markedly increased the yield of PGE2 at the expense of TxB2. Of the five morphologically discrete regions of the oviduct the vagina, infundibulum, and uterus contained the highest amounts of PGE and PGF, whereas the magnum and isthmus portions contain the least. TxB2 and 6-keto PGF1 alpha could not be detected in significant quantities in either region. These studies support the notion that PGE2 play a key role in the physiology of oviposition.

scholarly journals Influence of low- and high-protein diets on insulin and insulin-like growth factor-1 binding to skeletal muscle and liver in the growing rat

1991 ◽  
Vol 65 (1) ◽  
pp. 47-60 ◽  
Author(s):  
D. Dardevet ◽  
M. Manin ◽  
M. Balage ◽  
C. Sornet ◽  
J. Grizard
Keyword(s):  
Skeletal Muscle ◽  
Growth Factor ◽  
Dietary Protein ◽  
Binding Capacity ◽  
Plasma Concentrations ◽  
Insulin Binding ◽  
Metabolic Adaptation ◽  
Scatchard Analysis ◽  
Insulin Like Growth Factor ◽  
Hormone Binding

The influence of protein content of the diet on the plasma concentrations and binding to skeletal muscle and liver of insulin and insulin-like growth factor-1 (IGF-1), was studied in growing rats. Animals with a starting body-weight of 80 g received for an 11 d period isoenergetic diets containing (g/kg dry matter) 155 protein as controls (MP), or 55 (LP) or 300 (HP) protein. Food was offered as six equal meals/d. Daily food intakes provided adequate amounts of energy. Total plasma IGF-1 increased linearly as a function of dietary protein intake. Plasma insulin was lower in the LP than in the MP and HP groups. Hormone binding was studied in wheat-germ agglutinin (WGA) partially purified skeletal muscle receptor preparations. Each 125I-labelled hormone binding was competed for by increasing amounts of homologous and heterologous unlabelled hormone; this displacement needed lower concentrations of homologous than heterologous hormone. When compared with MP-diet feeding, the LP diet resulted in an increased ligand concentration for half-maximal binding. In addition the specific 125I-labelled insulin and 125I-labelled IGF-1 binding increased at all hormone concentrations and, as revealed by Scatchard analysis, the hormone binding capacity also rose (only significant for low-affinity insulin receptors and high-affinity IGF-1 receptors). The HP diet had little effect on hormone binding, except to increase insulin binding at very low insulin concentrations. Hormone binding was further studied in WGA partially purified liver receptor preparations. Those preparations did not exhibit any detectable specific 125I-labelled IGF-1 binding. The specific 125I-labelled insulin binding was not altered by dietary protein level. It is concluded that the increase in skeletal muscle insulin and IGF-1 binding along with a decrease in insulin and IGF-1 in the blood from rats fed on the LP diet, is consistent with the concept of an inverse relationship between plasma hormone and hormone binding. The physiological significance with respect to metabolic adaptation of muscle remains to be established

Anti-TSH receptor antibodies in Graves' disease modulate the kinetic behaviour of autologous thyroid TSH receptors. Evidence of the IgG-induced cross-linkage of autologous TSH receptors

1984 ◽  
Vol 105 (3) ◽  
pp. 330-340 ◽  
Author(s):  
Tjerk W. A. de Bruin ◽  
Daan van der Heide ◽  
Maria C. Krol
Keyword(s):  
Binding Sites ◽  
Plasma Membranes ◽  
Tsh Receptor ◽  
Human Thyroid ◽  
Graves Disease ◽  
Kinetic Behaviour ◽  
High Affinity ◽  
Cross Linkage ◽  
Receptor Antibodies ◽  
Tsh Receptor Antibodies

Abstract. The effect of the anti-TSH receptor antibodies present in the sera of 8 patients with Graves' disease on the affinity constant (Ka) and the number (R) of TSH receptors in autologous human thyroid plasma membranes was investigated. Kinetic analysis of [125I]bTSH binding to human thyroid plasma membranes in the presence of autologous Graves' and normal gammaglobulins was carried out by means of a computer fitting programme. Analysis of the TSH-TSH receptor interaction in the presence of TSH alone yielded curvilinear Scatchard plots, indicating the existence of two independent classes of binding sites (high affinity Ka: 8.5 ± 4.8 × 108 m−1; low affinity Ka: 5.3 ± 2.7 × 106 m−1). Similarly the Scatchard plot for this interaction in the presence of normal gammaglobulins is also curvilinear. Linear Scatchard plots, indicating the existence of only one class of high affinity TSH binding sites (Ka: 3.5 ± 1.8 × 108 m−1), were obtained for both autologous gammaglobulins and pure IgG from 8 patients with Graves' disease. The number of high affinity TSH binding sites in the presence of Graves' gammaglobulins had increased on the average by a factor 3.76 ± 0.74 (sd) with respect to the number found in the presence of normal gammaglobulins. This marked change in the kinetic behaviour of the TSH binding sites provided evidence that there is a direct interaction between anti-TSH receptor antibodies and autologous TSH receptors. Divalency of Graves' IgG or linkage of Fab fragments by anti-Fab antiserum proved to be necessary to produce this specific change in the kinetic behaviour of TSH binding sites. Graves' IgG monovalent Fab and Fc fragments had no effect. We suggest that the mechanism by which anti-TSH receptor antibodies in Graves' disease mimick the biological action of TSH is the IgG-induced cross-linkage of TSH receptors.

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