Journal articles on the topic 'Receptor affinity'
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1
Bailon, P., and D. V. Weber. "Receptor-affinity chromatography." Nature 335, no. 6193 (October 1988): 839–40. http://dx.doi.org/10.1038/335839a0.
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KOHANSKI, RONALD A., and M. DANIEL LANE. "Receptor Affinity Chromatography." Annals of the New York Academy of Sciences 447, no. 1 Biotin (June 1985): 373–85. http://dx.doi.org/10.1111/j.1749-6632.1985.tb18452.x.
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Dunn, W. A., T. P. Connolly, and A. L. Hubbard. "Receptor-mediated endocytosis of epidermal growth factor by rat hepatocytes: receptor pathway." Journal of Cell Biology 102, no. 1 (January 1, 1986): 24–36. http://dx.doi.org/10.1083/jcb.102.1.24.
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Substantial amounts of epidermal growth factor (EGF) are cleared from the circulation by hepatocytes via receptor-mediated endocytosis and subsequently degraded within lysosomes. We have used a combined biochemical and morphological approach to examine the fate of the receptor after exposure to EGF. Polyclonal antibodies were prepared against the purified receptor and their specificity established by immunoprecipitation and immunoblotting techniques. The EGF receptor was then localized by immunofluorescence and immunoperoxidase techniques and quantified on immunoblots. In untreated livers, EGF receptor was restricted to the sinusoidal and lateral surfaces of hepatocytes. 2-4 min after exposure of cells to EGF, the receptor was found in small vesicles (i.e., coated vesicles) as well as larger vesicles and tubules at the cell periphery. By 15 min the receptor was found in multivesicular endosomes located near bile canaliculi. Exposure of hepatocytes to EGF also resulted in a rapid loss of receptor protein from total liver homogenates and a decrease in its half-life from 8.7 h in control livers to 2.5 h. This EGF-induced loss of receptors was not observed when lysosomal proteinases were inhibited by leupeptin or when endosome/lysosome fusion was prevented by low temperature (16 degrees C). In the presence of leupeptin, receptor could be detected in structures identified as lysosomes using acid-phosphatase cytochemistry. All these results suggested rapid internalization of EGF receptors in response to ligand and degradation within lysosomes. However, four times more ligand was degraded at 8 h than the number of high-affinity (Kd of 8-15 nM) EGF-binding sites lost, suggesting either (a) high-affinity receptors were recycled, and/or (b) more than 300,000 receptors were available for EGF uptake. We identified and characterized a latent pool of approximately 300,000 low-affinity receptors (Kd approximately 200 nM) that could be separated on sucrose gradients from the plasma membrane pool of approximately 300,000 high-affinity receptors (Kd of 8-15 nM). Despite the differences in their binding affinities, the high- and low-affinity receptors appeared to be structurally identical and were both EGF-dependent protein kinases. In addition, the dynamics of the low-affinity receptors were consistent with a functional role in EGF uptake and delivery to lysosomes.
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Feldman, R. D., G. W. Hunninghake, and W. L. McArdle. "Beta-adrenergic-receptor-mediated suppression of interleukin 2 receptors in human lymphocytes." Journal of Immunology 139, no. 10 (November 15, 1987): 3355–59. http://dx.doi.org/10.4049/jimmunol.139.10.3355.
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Abstract Adrenergic receptor agonists are known to attenuate the proliferative response of human lymphocytes after activation; however, their mechanism of action is unknown. Since expression of interleukin 2 (IL-2) receptors is a prerequisite for proliferation, the effect of beta-adrenergic receptor agonists on lymphocyte IL-2 receptors was studied on both mitogen-stimulated lymphocytes and IL-2-dependent T lymphocyte cell lines. In both cell types the beta-adrenergic receptor agonist isoproterenol blocked the expression of IL-2 receptors, as determined with the IL-2 receptor anti-TAC antibody. To determine the effect of beta-adrenergic agonists on expression of the high affinity IL-2 receptors, [125I]IL-2 binding studies were performed at concentrations selective for high affinity sites. No significant effect of beta-adrenergic agonists on high affinity IL-2 receptor sites could be detected. The data demonstrate that beta-adrenergic receptor agonists down-regulate IL-2 receptors primarily affecting low affinity sites.
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Jonas, H. A., and L. C. Harrison. "Disulphide reduction alters the immunoreactivity and increases the affinity of insulin-like growth-factor-I receptors in human placenta." Biochemical Journal 236, no. 2 (June 1, 1986): 417–23. http://dx.doi.org/10.1042/bj2360417.
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We previously identified two forms of the insulin-like growth-factor-I (IGF-I) receptor in human placenta: a lower-affinity form reactive with an autoantiserum (B-2) to the insulin receptor and a higher-affinity non-immunoreactive form [Jonas & Harrison (1985) J. Biol. Chem. 260, 2288-2294]. Evidence is now presented that the lower-affinity immunoreactive forms are convertible into higher-affinity non-immunoreactive forms via reduction of receptor disulphide bonds. Treatment of placental membranes with increasing concentrations of dithiothreitol (DTT): (1) converted native Mr-290 000 heterotetrameric IGF-I receptors into Mr-180 000 dimers (determined by chemical cross-linking of 125I-IGF-I with disuccinimidyl suberate); (2) increased 125I-IGF-I binding, owing to an increase in receptor affinity; and (3) abolished the reactivity of Triton-solubilized IGF-I receptors with antiserum B-2 and transformed the curvilinear plot of IGF-I binding to a linear form. In isolated complexes between receptor and B-2 antibody, DTT increased 125I-IGF-I binding and released a single class of higher affinity IGF-I receptors of Mr 180,000. Thus DTT-treated IGF-I receptors have similar properties to the higher-affinity non-immunoreactive forms of the native receptor, except that reduced dimeric forms are not detected by cross-linking of 125I-IGF-I to native membranes. Cleavage of the inter-dimeric disulphide bonds is therefore not a prerequisite for higher-affinity binding or loss of immunoreactivity. These observations suggest that the thiol redox state of the IGF-I receptor in vivo is an important determinant of receptor conformation and therefore of the biological responses to IGF-I.
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Kristensen, C., A. S. Andersen, M. Hach, F. C. Wiberg, L. Schäffer, and T. Kjeldsen. "A single-chain insulin-like growth factor I/insulin hybrid binds with high affinity to the insulin receptor." Biochemical Journal 305, no. 3 (February 1, 1995): 981–86. http://dx.doi.org/10.1042/bj3050981.
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1. To investigate the structure/function relationship of the interaction between ligand and receptor in the insulin-like growth factor I (IGF-I) and insulin receptor systems we have prepared and characterized a single-chain insulin/IGF-I hybrid. The single-chain hybrid consists of the insulin molecule combined with the C domain of IGF-I. The single-chain hybrid was found to bind with high affinity to both truncated soluble insulin receptors and membrane-bound holoreceptors. The affinity for interacting with the soluble truncated insulin receptors was 55-94% relative to insulin, and affinity for membrane-bound insulin receptors was 113% of that of insulin. Furthermore we found that the affinity of the single-chain hybrid molecule for IGF-I receptors was 19-28% relative to IGF-I. 2. The affinity of the single-chain hybrid for chimeric insulin/IGF-I receptors exceeded that of either natural ligand. This indicates that coordinately changing domains of the receptors and the ligands can induce higher affinity of ligand for receptor, supporting the idea that these receptors have a common ligand-binding site [Kjeldsen, Andersen, Wiberg, Rasmussen, Schäffer, Balschmidt, Møller and Møller (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 4404-4408]. 3. In contrast with what was generally assumed about the ligand structure required for binding to the insulin receptor we demonstrate the first single-chain insulin analogue that can bind with high affinity to the insulin receptor.
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Dormann, Dirk, Ji-Yun Kim, Peter N. Devreotes, and Cornelis J. Weijer. "cAMP receptor affinity controls wave dynamics, geometry and morphogenesis inDictyostelium." Journal of Cell Science 114, no. 13 (July 1, 2001): 2513–23. http://dx.doi.org/10.1242/jcs.114.13.2513.
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Serpentine G-protein-coupled cAMP receptors are key components in the detection and relay of the extracellular cAMP waves that control chemotactic cell movement during Dictyostelium development. During development the cells sequentially express four closely related cAMP receptors of decreasing affinity. In this study, we investigated the effect of cAMP receptor type and affinity on the dynamics of cell-cell signalling in vivo, by measuring the dynamics of wave initiation and propagation in a variety of cAMP receptor mutants. We found that receptor affinity controls the frequency of wave initiation, but it does not determine wave propagation velocity, thus resulting in dramatic changes in wave geometry. In the limiting case, the affinity of the receptor is so low that waves can still be initiated but no stable centres form - thus, the cells cannot aggregate. In mounds, expression of low affinity receptors results in slow concentric waves instead of the normally observed multi-armed spiral waves. Under these conditions there is no rotational cell movement and the hemispherical mounds cannot transform into slugs. These results highlight the importance of receptor number and affinity in the proper control of cell-cell signalling dynamics required for the successful completion of development.
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Bevan, John A., Rosemary D. Bevan, and S. Martin Shreeve. "Variable receptor affinity hypothesis." FASEB Journal 3, no. 6 (April 1989): 1696–704. http://dx.doi.org/10.1096/fasebj.3.6.2564831.
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Yamashina, Masahiro, Takahiro Tsutsui, Yoshihisa Sei, Munetaka Akita, and Michito Yoshizawa. "A polyaromatic receptor with high androgen affinity." Science Advances 5, no. 4 (April 2019): eaav3179. http://dx.doi.org/10.1126/sciadv.aav3179.
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Biological receptors distinguish and bind steroid sex hormones, e.g., androgen-, progestogen-, and estrogen-type hormones, with high selectivity. To date, artificial molecular receptors have been unable to discriminate between these classes of biosubstrates. Here, we report that an artificial polyaromatic receptor preferentially binds a single molecule of androgenic hormones, known as “male” hormones (indicated with m), over progestogens and estrogens, known as “female” hormones (indicated with f), in water. Competitive experiments established the binding selectivity of the synthetic receptor for various sex hormones to be testosterone (m) > androsterone (m) >> progesterone (f) > β-estradiol (f) > pregnenolone (f) > estriol (f). These bindings are driven by the hydrophobic effect, and the observed selectivity arises from multiple CH-π contacts and hydrogen-bonding interactions in the semirigid polyaromatic cavity. Furthermore, micromolar fluorescence detection of androgen was demonstrated using the receptor containing a fluorescent dye in water.
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Broudy, Virginia C., Nancy L. Lin, Diana F. Sabath, Thalia Papayannopoulou, and Kenneth Kaushansky. "Human Platelets Display High-Affinity Receptors for Thrombopoietin." Blood 89, no. 6 (March 15, 1997): 1896–904. http://dx.doi.org/10.1182/blood.v89.6.1896.
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Abstract Thrombopoietin (Tpo) is a major regulator of megakaryopoiesis both in vivo and in vitro. Tpo initiates its biologic effects by binding to the Mpl receptor, which is a member of the hematopoietin receptor family. To define the Tpo binding characteristics of the Mpl receptor, we iodinated purified 70-kD recombinant human Tpo using the Bolton-Hunter reagent. Autoradiographic analysis of 125I-Tpo binding to normal human marrow mononuclear cells showed many grains specifically associated with megakaryocytes; there were no grains specifically associated with myeloblasts or erythroblasts. Equilibrium binding experiments with 125I-Tpo and normal human platelets showed a single class of high-affinity receptors (kd, 190 pmol/L) with approximately 30 Mpl receptors per platelet. Affinity cross-linking with 125I-Tpo showed that the Mpl receptor on platelets is of molecular weight ∼98 kD. Despite their sequence similarity, erythropoietin and Tpo did not cross-compete for binding to BaF3 cells engineered to coexpress Mpl receptor and erythropoietin receptor. Progeny of normal human burst-forming units-erythroid (BFU-E) contained Mpl receptor mRNA, and flow cytometric analysis showed the presence of Mpl receptor protein on the surface of these cells. These data indicate that display of the Mpl receptor is not limited to the megakaryocytic lineage, but also includes progeny of BFU-E. Like receptors for other hematopoietic cytokines, the binding affinity of the Mpl receptor for Tpo is high, with relatively few receptors displayed per cell. These results suggest that the effects of Tpo to speed red blood cell recovery after myelosuppressive therapy in vivo and to enhance colony-forming unit-erythroid generation in vitro may be mediated by direct interaction of Tpo and erythroid progenitor cells.
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Bradford, C. Samuel, Eliza A. Walthers, Brian T. Searcy, and Frank L. Moore. "Cloning, heterologous expression and pharmacological characterization of a kappa opioid receptor from the brain of the rough-skinned newt, Taricha granulosa." Journal of Molecular Endocrinology 34, no. 3 (June 2005): 809–23. http://dx.doi.org/10.1677/jme.1.01711.
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A full-length cDNA that encodes a kappa (κ) opioid receptor has been isolated from the brain of a urodele amphibian, the rough-skinned newt Taricha granulosa. The deduced protein contains 385 amino acids and possesses features commonly attributed to G protein-coupled receptors, such as seven putative transmembrane domains. The newt κ receptor has 75% sequence identity to κ opioid receptors cloned from mammals, and 66% sequence identity to the κ opioid receptor reported for the zebrafish, with the greatest divergence in the extracellular N-terminus, the second and third extracellular loops and the intracellular C-terminus. Membranes isolated from COS-7 cells expressing the newt κ receptor possessed a single, high-affinity (Kd =1.5 nM) binding site for the κ-selective agonist U69593. In competition binding assays, the expressed newt κ receptor displayed high affinity for the κ-selective agonists GR89696, dynorphin A(1–13), U69593, U50488 and BRL52537, as well as the κ-selective antagonist nor-binaltorphimine and the non-selective antagonist naloxone. Rank order potencies and affinity constants were similar in competition binding studies that used either whole brain homogenates or membranes isolated from COS-7 cells expressing the newt κ receptor. The expressed receptor displayed essentially no affinity for the δ-selective agonist DPDPE ([d-penicillamine, d-penicillamine]enkephalin), but showed moderate affinity for the μ-selective agonist DAMGO ([d-Ala-MePhe, Gly-ol]enkephalin) and moderately high affinity for nociceptin (orphanin FQ), the endogenous ligand for the opioid receptor-like (ORL)1 receptor. These findings support the conclusions that a gene for the κ opioid receptor is expressed in amphibians and that the pharmacology of the newt κ receptor closely matches mammalian κ opioid receptors. However, the functional dichotomy between the classic opioid receptors (κ, δ, μ) and ORL1 appears less strict in amphibians than in mammals.
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Knol, Remco J. J., Jan C. van den Bos, Anton G. M. Janssen, Kora de Bruin, Berthe L. F. van Eck-Smit, and Jan Booij. "Synthesis and In Vitro Evaluation of Novel Nortropane Derivatives as Potential Radiotracers for Muscarinic M2 Receptors." International Journal of Molecular Imaging 2011 (June 13, 2011): 1–7. http://dx.doi.org/10.1155/2011/709416.
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Disturbances of the cerebral cholinergic neurotransmitter system are present in neurodegenerative disorders. SPECT or PET imaging, using radiotracers that selectively target muscarinic receptor subtypes, may be of value for in vivo evaluation of such conditions. 6β-acetoxynortropane, a potent muscarinic M2 receptor agonist, has previously demonstrated nanomolar affinity and high selectivity for this receptor. Based on this compound we synthesized four nortropane derivatives that are potentially suitable for SPECT imaging of the M2 receptor. 6β-acetoxynortropane and the novel derivatives were tested in vitro for affinity to the muscarinic M1−3 receptors. The original 6β-acetoxynortropane displayed high affinity (Ki=70–90 nM) to M2 receptors and showed good selectivity ratios to the M1 (65-fold ratio) and the M3 (70-fold ratio) receptors. All new derivatives showed reduced affinity to the M2 subtype and loss of subtype selectivity. It is therefore concluded that the newly synthesized derivatives are not suitable for human SPECT imaging of M2 receptors.
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Jong, Y. J., L. R. Dalemar, B. Wilhelm, and N. L. Baenziger. "Human bradykinin B2 receptors isolated by receptor-specific monoclonal antibodies are tyrosine phosphorylated." Proceedings of the National Academy of Sciences 90, no. 23 (December 1, 1993): 10994–98. http://dx.doi.org/10.1073/pnas.90.23.10994.
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We report the immunoaffinity isolation of bradykinin B2 receptors in a tyrosine-phosphorylated state from WI-38 human lung fibroblasts. We generated six monoclonal antibodies directed against B2 bradykinin receptor biologic activity mediating prostaglandin E2 production in WI-38. These cells express a repertoire of bradykinin receptor affinity forms with closely correlated biologic activity and [3H]bradykinin binding. Some of the monoclonal antibodies selectively recognize intermediate-affinity (Kd = 5.6 nM) or low-affinity (Kd = 42 nM) receptor forms, whereas others recognize epitopes common to both. The monoclonal antibodies block bradykinin binding and biologic activity. Immunoaffinity chromatography on an immobilized monoclonal antibody of intermediate- plus low-affinity specificity yields WI-38 B2 receptors with intact [3H]bradykinin binding activity and a molecular mass of 78 kDa. The same band is immunoblotted by all the monoclonal antibodies, indicating a similar molecular mass for receptor forms of different affinity. Anti-phosphotyrosine antibodies demonstrate that the receptors are tyrosine phosphorylated, with implications for receptor function and regulation. Genistein completely inhibits bradykinin-mediated prostaglandin E2 production with an IC50 of 8 microM, indicating that tyrosine kinase activity is critical for the signal transduction leading to arachidonic acid release.
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Robb, R. J., and C. M. Rusk. "High and low affinity receptors for interleukin 2: implications of pronase, phorbol ester, and cell membrane studies upon the basis for differential ligand affinities." Journal of Immunology 137, no. 1 (July 1, 1986): 142–49. http://dx.doi.org/10.4049/jimmunol.137.1.142.
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Abstract Cellular binding sites for IL 2 exist in two forms which differ with respect to their apparent affinity for the factor. The present studies were designed to evaluate various models for the difference. Receptor-mediated internalization and covalent receptor-ligand coupling were discounted as explanations on the basis of ligand binding and elution studies on permeabilized cells and cell membranes. Phosphorylation of the receptor during activation of protein kinase C failed to modulate the ratio of high and low affinity sites, demonstrating that it also did not provide a potential mechanism. Selective destruction of low affinity receptors with pronase, on the other hand, indicated that the two forms of binding sites differed significantly in their cell surface structure. Either the two types of receptor consist of distinct molecules or the conformation of the high affinity binding sites renders them more resistant to proteolysis. Antibody inhibition studies revealed that the high affinity receptors remaining after protease treatment and their low affinity counterparts both utilized the same ligand-binding component. Thus, this result ruled out the possibility of two totally distinct receptor structures. Together, the findings support the hypothesis that other membrane components modify the conformation of the ligand-binding polypeptide to confer a high affinity protease-resistant configuration.
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Walker, F., and A. W. Burgess. "Transmodulation of the epidermal-growth-factor receptor in permeabilized 3T3 cells." Biochemical Journal 256, no. 1 (November 15, 1988): 109–15. http://dx.doi.org/10.1042/bj2560109.
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Binding of murine epidermal growth factor (EGF) to its high-affinity receptor can be modulated by a variety of structurally unrelated mitogens. The transmodulation, however, is temperature-dependent and has not been observed in isolated membranes. We report here the transmodulation of high-affinity EGF receptors by platelet-derived growth factors (PDGF) and tumour-promoting phorbol esters in 3T3 cells even when they are rendered incapable of fluid-phase endocytosis by treatment with phenylarsine oxide or by permeabilization with lysophosphatidylcholine. The relative affinity of the EGF receptors in the absence of modulating agents is not significantly altered by phenylarsine oxide treatment. Thus the difference in affinity between the two classes of EGF receptors seems to be unrelated to dynamic membrane changes or to differential rates of internalization. In permeabilized cells, non-hydrolysable GTP analogues transmodulate the high-affinity EGF receptor; however, the effects of these analogues are blocked by the protein kinase C inhibitor chlorpromazine. In contrast, transmodulation by PDGF is not blocked by chloropromazine. Thus the high-affinity EGF receptor can be transmodulated by both protein kinase C-dependent or -independent pathways, and the transmodulation processes do not require fluid-phase endocytosis.
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Ortega, Gabriel, Davide Mariottini, Alessandra Troina, Frederick W. Dahlquist, Francesco Ricci, and Kevin W. Plaxco. "Rational design to control the trade-off between receptor affinity and cooperativity." Proceedings of the National Academy of Sciences 117, no. 32 (July 29, 2020): 19136–40. http://dx.doi.org/10.1073/pnas.2006254117.
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Cooperativity enhances the responsiveness of biomolecular receptors to small changes in the concentration of their target ligand, albeit with a concomitant reduction in affinity. The binding midpoint of a two-site receptor with a Hill coefficient of 1.9, for example, must be at least 19 times higher than the dissociation constant of the higher affinity of its two binding sites. This trade-off can be overcome, however, by the extra binding energy provided by the addition of more binding sites, which can be used to achieve highly cooperative receptors that still retain high affinity. Exploring this experimentally, we have employed an “intrinsic disorder” mechanism to design two cooperative, three-binding-site receptors starting from a single-site—and thus noncooperative—doxorubicin-binding aptamer. The first receptor follows a binding energy landscape that partitions the energy provided by the additional binding event to favor affinity, achieving a Hill coefficient of 1.9 but affinity within a factor of 2 of the parent aptamer. The binding energy landscape of the second receptor, in contrast, partitions more of this energy toward cooperativity, achieving a Hill coefficient of 2.3, but at the cost of 4-fold poorer affinity than that of the parent aptamer. The switch between these two behaviors is driven primarily by the affinity of the receptors’ second binding event, which serves as an allosteric “gatekeeper” defining the extent to which the system is weighted toward higher cooperativity or higher affinity.
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Finbloom, D. S., and L. M. Wahl. "Characterization of a novel low affinity receptor for IFN-gamma on adherent human monocytes by radioligand binding studies and chemical cross-linking." Journal of Immunology 142, no. 7 (April 1, 1989): 2314–20. http://dx.doi.org/10.4049/jimmunol.142.7.2314.
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Abstract Freshly isolated monocytes in suspension express 2000 to 4000 high affinity receptors for IFN-gamma. Because monocytes change phenotypically as they migrate out of the circulation and adhere to extracellular matrix, modulation of the expression of IFN-gamma receptors may occur. In order to determine if adherence alone modulates the receptor for IFN-gamma, we have studied receptor expression in adherent human peripheral blood monocytes. Elutriation-purified monocytes were allowed to adhere to polystyrene overnight at 37 degrees C. These cells now expressed 1 to 2 x 10(5) low affinity (Ka = 10(8) liters/M) receptors for [125I]rIFN-gamma. Binding to this receptor was specific and saturable. The expression of these receptors occurred rapidly (within 3 h) after adherence and was not inhibited by cycloheximide treatment. Binding to the receptor was abrogated by treating cells with trypsin, but was enhanced after treatment with alkaline protease or proteinase K. mAb against the high affinity receptor did not block binding to the low affinity receptor on adherent cells. The low affinity receptor transduced a signal to the cell as measured by the IFN-gamma-induced enhancement in FcR for human IgG1. The structure of the receptor on adherent cells was investigated by chemical cross-linking techniques. A receptor-[125I]rIFN-gamma complex was observed by SDS-PAGE to have a Mr of 180,000 to 200,000. Reduction of this complex with 2-ME resulted in the loss of the high Mr complex and the appearance of a doublet of lower Mr of 68,000 and 82,000. In contrast, cross-linking of monocytes in suspension yielded a complex of 110,000 to 120,000 Mr, which was unchanged upon reduction. Upon adherence, human monocytes express large numbers of a novel receptor for rIFN-gamma which is capable of stimulating the cell. This receptor appears to be composed of at least two components which are disulfide linked and structurally differs from the high affinity receptor on nonadherent monocytes.
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Yeung, S. M. H., E. Perez-Reyes, and D. M. F. Cooper. "Hydrodynamic properties of adenosine Ri receptors solubilized from rat cerebral-cortical membranes." Biochemical Journal 248, no. 3 (December 15, 1987): 635–42. http://dx.doi.org/10.1042/bj2480635.
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Adenosine Ri receptors and inhibitory guanine-nucleotide-regulatory components were solubilized from rat cerebral-cortical membranes with sodium cholate. (-)-N6-Phenylisopropyl[2,8-3H]adenosine [(3H]PIA) binds with high affinity to the soluble receptors, which retain the pharmacological specificity of adenosine Ri receptors observed in membranes. The binding is regulated by bivalent cations and guanine nucleotides. Bivalent cations increase [3H]PIA binding by increasing both the affinity and the apparent number of receptors. Guanine nucleotides decrease agonist binding by increasing the dissociation of the ligand-receptor complex. Adenosine agonists stabilize the high-affinity form of the soluble receptor. The hydrodynamic properties of the adenosine receptor were determined with cholate extracts of membranes that were treated with [3H]PIA. Sucrose-gradient-centrifugation analysis indicates that the receptor has a sedimentation coefficient of 7.7 S. The receptor is eluted from Sepharose 6B columns with an apparent Stokes radius of 7.2 nm. Labelling of either sucrose-gradient or gel-filtration-column fractions with pertussis toxin and [32P]-NAD+ reveals that both the 41,000- and 39,000-Mr substrates overlap with the receptor activity. These studies suggest that the high-affinity adenosine-receptor-binding activity in the cholate extract represents a stable R1-N complex.
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Soraya, hiva, Ruohollah Seddigh, Fatemeh Hadi, and Mohammad Faramarzi. "Chemical cannabis; The New Trend of addiction in Iran." Iranian Journal of Psychiatry and Clinical Psychology 28, no. 1 (April 20, 2022): 10. http://dx.doi.org/10.32598/ijpcp.28.1.4010.1.
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Synthetic cannabinoids (SC) are a heterogeneous group of substances with a high affinity for cannabinoid receptors. Unlike Δ9-tetrahydrocannabinol (THC), synthetic cannabinoids are incredibly potent, highly productive, have more affinity for the Cannabinoid receptor type 1 (CB1), and Cannabinoid receptor type 2 (CB2), and are designed to accelerate the effects of tetrahydrocannabinol. Also, there is experimental evidence that SCs acts on non-cannabinoid receptors, such as the 5-HT2B receptor or dopaminergic receptors. (1, 2).
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McArthur, K. E., C. L. Wood, M. S. O'Dorisio, Z. C. Zhou, J. D. Gardner, and R. T. Jensen. "Characterization of receptors for VIP on pancreatic acinar cell plasma membranes using covalent cross-linking." American Journal of Physiology-Gastrointestinal and Liver Physiology 252, no. 3 (March 1, 1987): G404—G412. http://dx.doi.org/10.1152/ajpgi.1987.252.3.g404.
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Vasoactive intestinal peptide (VIP) receptors on guinea pig pancreatic acini differ from those on all other tissues in containing a high-affinity VIP receptor and a low-affinity VIP receptor that has a high affinity for secretin. To characterize the molecular components of these receptors, 125I-VIP was covalently cross-linked to these receptors by four different cross-linking agents: disuccinimidyl suberate, ethylene glycol bis (succinimidyl succinate), dithiobis (succinimidylpropionate), and m-maleimidobenzoyl N-hydroxysuccinimide ester. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated a single major polypeptide band of Mr 45,000 and a minor polypeptide band of Mr 30,000 were cross-linked to 125I-VIP. Covalent cross-linking only occurred when a cross-linking agent was added, was inhibited by GTP, was inhibited by VIP receptor agonists or antagonists that interact with VIP receptors, and not by other pancreatic secretagogues that interact with different receptors. For inhibiting both cross-linking and binding of 125I-VIP to the major polypeptide Mr 45,000 and the minor polypeptide Mr 30,000 components, the relative potencies were VIP greater than helodermin greater than rat growth hormone releasing factor greater than peptide histidine isoleucine greater than secretin. The apparent molecular weight of the cross-linked polypeptides were unchanged by dithiothreitol. Thus the high-affinity VIP receptor on pancreatic acinar cell membranes consists of a single major polypeptide of Mr 45,000, and this polypeptide is not a subunit of a larger disulfide-linked structure. Furthermore, either the low-affinity VIP/secretin-preferring receptor was not covalently cross-linked under the experimental conditions or it consists of a major polypeptide with the same molecular weight as the high-affinity VIP receptor.
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Chatterjee, Tapan K., and Rory A. Fisher. "Multiple affinity and guanine nucleotide sensitive forms of the calcitonin gene related peptide (CGRP) receptor." Canadian Journal of Physiology and Pharmacology 73, no. 7 (July 1, 1995): 968–73. http://dx.doi.org/10.1139/y95-134.
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Calcitonin gene related peptide (CGRP) is a novel neuropeptide with an impressive array of biological actions consistent with its diverse tissue distribution and suggested role as neurotransmitter or neuromodulator. Binding sites for CGRP with properties consistent with those of receptors are present in both central and peripheral tissues. Radioligand binding studies were performed to investigate the fundamental processes underlying CGRP receptor activation and signaling following agonist occupancy of the receptor. These studies documented the existence of a selective, high affinity, and homogeneous population of binding sites for CGRP in membranes prepared from central and various peripheral tissues. The affinity of [125I]CGRP for these sites was regulated by GTP or its stable analog GTPγS, indicating coupling of CGRP receptors to G-protein(s). Kinetic studies documented the existence of the CGRP receptor in multiple affinity states when both coupled to and uncoupled from G-proteins(s). These findings suggest that CGRP occupancy of its receptor induces conformational changes in the receptor that may be involved in its coupling to G-proteins and that the resulting ligand–receptor–G-protein ternary complex exists in multiple affinity conformational states. It seems likely that the multiple affinity states of the CGRP receptor ternary complex are involved differentially in signaling by and desensitization of the receptor. This evidence for agonist-induced conformational changes in a G-protein-coupled receptor prior to its coupling with G-protein(s) and for the existence of the ligand–receptor–G-protein ternary complex in multiple affinity conformational states is novel and extends our current understanding of the nature of the processes involved in agonist-dependent activation of G-protein-coupled receptors.Key words: calcitonin gene related peptide receptor, G-protein, multiple affinity.
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Felder, S., J. LaVin, A. Ullrich, and J. Schlessinger. "Kinetics of binding, endocytosis, and recycling of EGF receptor mutants." Journal of Cell Biology 117, no. 1 (April 1, 1992): 203–12. http://dx.doi.org/10.1083/jcb.117.1.203.
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This report describes analysis of factors which regulate the binding of EGF to EGF receptor, receptor internalization, and receptor recycling. Three different methods were used to inhibit high-affinity EGF binding as measured at equilibrium: treatment of cells with an active phorbol ester (PMA), binding of a mAb directed against the EGF receptor (mAb108), and truncation of most of the cytoplasmic domain of the receptor. These treatments reduced the rate at which low concentrations of EGF bound to cells, but did not affect the rate of EGF dissociation. We conclude that high-affinity EGF binding on living cells results from a difference in the apparent on rate of EGF binding. We then used these conditions and cell lines to test for the rate of EGF internalization at different concentrations of EGF. We demonstrate that internalization of the EGF receptor is stimulated roughly 50-fold at saturating concentrations of EGF, but is stimulated an additional two- to threefold at low concentrations (less than 1 nM). Four treatments reduce the rate of internalization of low concentrations of EGF to the rate seen at saturating EGF concentrations. Phorbol ester treatment and mAb108 binding to "wild type" receptor reduce this rate (and reduce high-affinity binding). Point mutation at Lys721 (kinase negative EGF receptor) and point mutation at Thr654 (removing a major site of protein kinase C phosphorylation) reduce the internalization rate, without affecting high-affinity binding. We suggest that while EGF stimulates endocytosis for all receptors, high-affinity receptors bind and are internalized more quickly than low-affinity receptors. Tyrosine kinase activity and the Thr654 region appear necessary for this response.
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Jacobs, AM, and F. Youngblood. "Opioid receptor affinity for agonist-antagonist analgesics." Journal of the American Podiatric Medical Association 82, no. 10 (October 1, 1992): 520–24. http://dx.doi.org/10.7547/87507315-82-10-520.
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Analgesic medications are distributed to a variety of receptors within the central nervous system. Activity at these receptors (mu 1, mu, sigma, delta, kappa) results in both the beneficial pain-relieving effects of analgesics as well as undesirable side effects. The mixed agonist-antagonist class of analgesics offers the potential benefit of greater receptor site selectivity while diminishing the incidence of adverse sequelae, such as respiratory depression. Traditionally, it has been suggested that mixed agonist-antagonist medications may be associated with decreased analgesic effectiveness. However, newer agents of this mixed class may result in effective analgesia while diminishing the incidence of side effects.
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Tonkin, K. S., M. Berger, and M. Ormerod. "Epidermal growth factor receptor status in four carcinoma of the cervix cell lines." International Journal of Gynecologic Cancer 1, no. 4 (1991): 185–92. http://dx.doi.org/10.1046/j.1525-1438.1991.01040185.x.
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We have evaluated aspects of the EGF receptor content of four human cell lines derived from patients with previously untreated carcinoma of the cervix. Scatchard analysis revealed that three of the lines possessed approximately 2×105 low-affinity and 2×104 high-affinity receptors, whereas the fourth line had approximately 9×104 low-affinity receptors and 9×103 high-affinity receptors. Immunocytochemical staining using the monoclonal antibody EGFR1 showed wide intra- and inter-line variation in staining intensity. Flow cytometric analysis of EGFR1 demonstrated a fivefold difference in staining intensity between lines. Thirteen cloned derivatives of one of the lines exhibited a 200% variation in EGFR1 staining intensity. There were no differences in radiosensitivity in four of the cloned lines with different EGF receptor levels. Southern blotting analysis did not reveal any rearrangement or amplication of the EGF receptor gene. These three different methods for determining receptor content produced variations in the ranking of receptor number across the four cell lines. These studies with cervix carcinoma cell lines demonstrate the presence of varied levels of EGF receptors according to the methodology used. This may reflect differences in biological characteristics of the cell lines evaluated.
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Baum, C. M., J. P. McKearn, R. Riblet, and J. M. Davie. "Polymorphism of Fc receptor on murine B cells is Igh-linked." Journal of Experimental Medicine 162, no. 1 (July 1, 1985): 282–96. http://dx.doi.org/10.1084/jem.162.1.282.
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Analysis of mouse IgG binding to Fc receptors on mouse B cells indicates that the IgG1, IgG2a, and IgGb subclasses bind to the same receptor. No differences in affinity were detected among subclass or between mouse strains. This same receptor bound rat IgG with an affinity that differed between mouse strains. This polymorphism in affinity for rat IgG maps to chromosome 12 distal to the Igh locus.
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Mehta, S. R., S. R. Grant, and A. L. Maizel. "Characterization of the cell surface receptors for human B cell growth factor of 12,000 molecular weight." Journal of Immunology 137, no. 7 (October 1, 1986): 2210–14. http://dx.doi.org/10.4049/jimmunol.137.7.2210.
Full textAbstract:
Abstract Quiescent normal human B cells have been shown to require an activation step before proliferating in response to B cell growth factor (BCGF) of 12,000 m.w. (12 kd). One effect of cell activation has been the putative acquisition of specific cell surface growth factor receptors. In this report, the existence of such receptors has been confirmed by using purified radioiodinated BCGF-12 kd. BCGF-12 kd receptors on activated B cells have been shown to be distinct form those interacting with IL 2. Scatchard analysis revealed both high affinity receptor sites with an apparent Kd of 28.6 pM and low affinity receptor sites with Kd of 1.2 nM on freshly prepared, anti-IgM activated peripheral blood B cells. Human B cells grown in culture for extended periods of time in the presence of BCGF-12 kd also displayed high affinity receptor sites (Kd, 41.4 pM) and low affinity receptor sites (Kd, 0.9 nM). The action of BCGF-12 kd therefore appears to be mediated by binding to its lineage-specific receptors on the cell surface.
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Turner, AM, LG Bennett, NL Lin, J. Wypych, TD Bartley, RW Hunt, HL Atkins, KE Langley, V. Parker, and F. Martin. "Identification and characterization of a soluble c-kit receptor produced by human hematopoietic cell lines." Blood 85, no. 8 (April 15, 1995): 2052–58. http://dx.doi.org/10.1182/blood.v85.8.2052.bloodjournal8582052.
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Stem cell factor (SCF) triggers cell growth by binding to cell surface c-kit receptors. Soluble forms of several cytokine receptors have been described and may play a role in the modulation of cytokine activity in vivo. For these reasons, we investigated whether human hematopoietic cells produce soluble c-kit receptors. The human leukemia cell lines OCIM1 and MO7e display approximately 80,000 and approximately 35,000 high-affinity cell surface c-kit receptors, respectively. Soluble c-kit receptors were detected by enzyme immunoassay in OCIM1 and MO7e culture supernatants. We determined the molecular weight and binding affinity of soluble c-kit receptor produced by OCIM1 cells, soluble c-kit receptor purified from human serum, and recombinant soluble c-kit receptor expressed in CHO cells. The three soluble c-kit receptors each have a molecular weight of 98 kD. Quantitative binding experiments with 125I-SCF indicate that the soluble c-kit receptors obtained from human serum or OCIM1 cells have binding affinities for SCF of approximately 200 to 300 pmol/L, in contrast to the recombinant form, which has a binding affinity of approximately 1.5 nmol/L. All three forms of the soluble c-kit receptor were able to compete with c-kit receptors on OCIM1 cells for 125I-SCF binding. Thus human hematopoietic cells can produce a soluble form of the c-kit receptor that retains high-affinity SCF binding activity. We speculate that the soluble c-kit receptor may bind SCF and function as a receptor antagonist in vivo.
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Sawyer, ST, SB Krantz, and K. Sawada. "Receptors for erythropoietin in mouse and human erythroid cells and placenta." Blood 74, no. 1 (July 1, 1989): 103–9. http://dx.doi.org/10.1182/blood.v74.1.103.103.
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Abstract High and lower affinity receptors for erythropoietin (EP) were initially identified on a very pure population of EP-responsive erythroblasts obtained from the spleens of mice infected with anemia strain of Friend virus (FVA). The structure of the receptor for EP in these cells was determined to be proteins of 100 and 85 Kd by cross- linking 125I-EP. In this investigation, studies on the receptors for EP were extended to other mouse erythroid cells and human erythroid cells as well as to the placentas of mice and rats. Only lower affinity receptors for EP were detected on erythroblasts purified from the spleens of mice infected with the polycythemia strain of Friend virus and a murine erythroleukemia cell line, both of which are not responsive to EP in culture. Internalization of 125I-EP was observed in both groups of cells. The structure of the receptor determined by cross- linking 125I-EP was two equally labeled proteins of 100 Kd and 85 Kd molecular mass in all these mouse erythroid cells. The structure of the receptor was found to be very similar in human erythroid colony forming cells cultured from normal blood. These cells respond to EP with erythroid maturation and were previously shown to have high and lower affinity receptors. Placentas from mice and rats were found to have only lower affinity receptors for EP, and when placental membranes were cross-linked to 125I-EP, the same 100 Kd and 85 Kd bands were found as seen in mouse and human erythroid cells. The structure of the receptor was similar in cells that have high affinity receptors (FVA-infected and human erythroid colony-forming cells) and nonresponsive erythroid cells and placenta that have lower affinity receptors, but only the cells with the high affinity receptors respond to the addition of EP with erythroid maturation.
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Sawyer, ST, SB Krantz, and K. Sawada. "Receptors for erythropoietin in mouse and human erythroid cells and placenta." Blood 74, no. 1 (July 1, 1989): 103–9. http://dx.doi.org/10.1182/blood.v74.1.103.bloodjournal741103.
Full textAbstract:
High and lower affinity receptors for erythropoietin (EP) were initially identified on a very pure population of EP-responsive erythroblasts obtained from the spleens of mice infected with anemia strain of Friend virus (FVA). The structure of the receptor for EP in these cells was determined to be proteins of 100 and 85 Kd by cross- linking 125I-EP. In this investigation, studies on the receptors for EP were extended to other mouse erythroid cells and human erythroid cells as well as to the placentas of mice and rats. Only lower affinity receptors for EP were detected on erythroblasts purified from the spleens of mice infected with the polycythemia strain of Friend virus and a murine erythroleukemia cell line, both of which are not responsive to EP in culture. Internalization of 125I-EP was observed in both groups of cells. The structure of the receptor determined by cross- linking 125I-EP was two equally labeled proteins of 100 Kd and 85 Kd molecular mass in all these mouse erythroid cells. The structure of the receptor was found to be very similar in human erythroid colony forming cells cultured from normal blood. These cells respond to EP with erythroid maturation and were previously shown to have high and lower affinity receptors. Placentas from mice and rats were found to have only lower affinity receptors for EP, and when placental membranes were cross-linked to 125I-EP, the same 100 Kd and 85 Kd bands were found as seen in mouse and human erythroid cells. The structure of the receptor was similar in cells that have high affinity receptors (FVA-infected and human erythroid colony-forming cells) and nonresponsive erythroid cells and placenta that have lower affinity receptors, but only the cells with the high affinity receptors respond to the addition of EP with erythroid maturation.
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Bhuiyan, Mohiuddin Ahmed, Mohammad Shahriar, and Takafumi Nagatomo. "Binding Affinity of Candesartan, Losartan, Telmisartan and Valsartan with Angiotensin II Receptor 1 Subtype." Bangladesh Pharmaceutical Journal 16, no. 1 (April 7, 2013): 10–14. http://dx.doi.org/10.3329/bpj.v16i1.14484.
Full textAbstract:
This study was designed to examine the interaction in the binding of selective angiotensin II receptor antagonists towards angiotensin II type 1 receptor. The AT1 antagonists used in this study were valsartan, candesartan and losartan. Wild type AT1 receptors were transiently expressed in COS-7 cells and the expressed protein was isolated. The binding affinities of agonist and these four AT1 antagonists were determined towards AT1 receptors with the help of radioligand binding studies. The binding affinity of candesartan has been found to be maximum having a pKi value of 8.61±0.21 whereas losartan showed lowest binding affinity among the antagonists (pKi=7.17±0.07). Telmisartan also showed high (pKi=8.19±0.04) and valsartan had moderate binding affinity (pKi=7.65±0.12) towards AT1 receptors. The results of the study suggested that candesartan interacts very strongly with the receptor which is consistent with the maximum number of binding sites of in the chemical structure of candesartan. On the other hand, losartan has lower number of binding sites with the amino acid residues of AT1 receptor and as a result it showed the minimum affinity towards the receptor. DOI: http://dx.doi.org/10.3329/bpj.v16i1.14484 Bangladesh Pharmaceutical Journal 16(1): 10-14, 2013
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Faull, R. J., N. L. Kovach, J. M. Harlan, and M. H. Ginsberg. "Stimulation of integrin-mediated adhesion of T lymphocytes and monocytes: two mechanisms with divergent biological consequences." Journal of Experimental Medicine 179, no. 4 (April 1, 1994): 1307–16. http://dx.doi.org/10.1084/jem.179.4.1307.
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We show that the adhesion of T lymphoid cells to immobilized fibronectin can be increased by two distinct mechanisms. The first is by increasing the affinity of the fibronectin receptor/ligand interaction using the anti-beta 1 integrin monoclonal antibody 8A2. The second is by treating the cells with phorbol 12-myristate 13-acetate (PMA), which alters events that occur after receptor occupancy (e.g., cell spreading) without affecting receptor affinity. The effects of these two mechanisms on adhesion in the presence of physiological concentrations of soluble fibronectin suggest that they have different biological consequences. Under these conditions, the net effect of increasing the affinity of the fibronectin receptors is to decrease cell adhesion, whereas the increase in adhesion induced by PMA is unaffected. This suggests that the high affinity receptors are not primarily available for cell adhesion under these circumstances, and that they have an alternative function. We further show that high affinity binding of soluble fibronectin can be induced by either differentiation of the monocytic cell line THP-1 or by cross-linking the T cell receptor complexes on the T lymphoid cell line HUT-78. The differentiated monocytic cells express two populations of fibronectin receptors: a minority in a high affinity state, and the majority in a low affinity state. Thus they will both continue to adhere in the presence of physiological concentrations of soluble fibronectin and bind significant amounts of soluble fibronectin at the cell surface.
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Davison, E., I. Kirby, T. Elliott, and G. Santis. "The Human HLA-A*0201 Allele, Expressed in Hamster Cells, Is Not a High-Affinity Receptor for Adenovirus Type 5 Fiber." Journal of Virology 73, no. 5 (May 1, 1999): 4513–17. http://dx.doi.org/10.1128/jvi.73.5.4513-4517.1999.
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ABSTRACT The coxsackie B virus and adenovirus receptor (CAR) and the major histocompatibility complex (MHC) class I α2 domain have been identified as high-affinity cell receptors for adenovirus type 5 (Ad5) fiber. In this study we show that CAR but not MHC class I allele HLA-A*0201 binds to Ad5 with high affinity when expressed on hamster cells. When both receptors are coexpressed on the cell surface of hamster cells, Ad5 fiber bind to a single high-affinity receptor, which is CAR.
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Budel, LM, O. Elbaz, H. Hoogerbrugge, R. Delwel, LA Mahmoud, B. Lowenberg, and IP Touw. "Common binding structure for granulocyte macrophage colony-stimulating factor and interleukin-3 on human acute myeloid leukemia cells and monocytes." Blood 75, no. 7 (April 1, 1990): 1439–45. http://dx.doi.org/10.1182/blood.v75.7.1439.1439.
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Abstract Granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin-3 (IL-3) control the proliferation of human acute myeloid leukemia (AML) cells in vitro. Previously, we have shown that receptors for GM-CSF and IL-3 are often coexpressed on AML cells. Here we present experiments with purified AML blasts, normal monocytes, and granulocytes that were conducted to analyze the properties of GM-CSF and IL-3 binding proteins in more detail. On AML cells from eight cases we demonstrate two types of GM-CSF receptors: one with low affinity (dissociation constant [kd] 5.1 to 24.8 nmol/L) and one with a high affinity (kd 31 to 104 pmol/L). These AML cells also expressed high affinity receptors for IL-3 (kd 24 to 104 pmol/L). Cross-competition experiments showed that an excess concentration of nonlabeled IL-3 completely prevented the high affinity binding of radiolabled GM-CSF. This competition occurred at 37 degrees C as well as 4 degrees C. Low affinity GM-CSF binding was not affected by IL-3. Binding of radiolabeled IL-3 could be prevented by nonlabeled GM-CSF. In certain cases, this competition was complete, whereas in others only partial (49% to 77%) reduction of the radiolabeled IL-3 binding was seen. On the basis of these ligand binding features, we propose the existence of three receptor types on AML cells: (1) low affinity GM-CSF receptors that do not bind IL-3, (2) dual high affinity GM-CSF/IL-3 receptors, and (3) high affinity IL-3 receptors that do not bind GM-CSF. We could also demonstrate these receptor types on normal monocytes. In addition, a fourth type of receptor was apparent on normal granulocytes (4), incapable of binding IL-3 and with an intermediate affinity for GM-CSF (approximately 400 pmol/L). Chemical crosslinking showed that GM-CSF and IL-3 both bind to proteins with molecular weight values of 130, 105, and 75, which provides additional evidence for the existence of a common GM-CSF/IL-3 receptor complex.
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Budel, LM, O. Elbaz, H. Hoogerbrugge, R. Delwel, LA Mahmoud, B. Lowenberg, and IP Touw. "Common binding structure for granulocyte macrophage colony-stimulating factor and interleukin-3 on human acute myeloid leukemia cells and monocytes." Blood 75, no. 7 (April 1, 1990): 1439–45. http://dx.doi.org/10.1182/blood.v75.7.1439.bloodjournal7571439.
Full textAbstract:
Granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin-3 (IL-3) control the proliferation of human acute myeloid leukemia (AML) cells in vitro. Previously, we have shown that receptors for GM-CSF and IL-3 are often coexpressed on AML cells. Here we present experiments with purified AML blasts, normal monocytes, and granulocytes that were conducted to analyze the properties of GM-CSF and IL-3 binding proteins in more detail. On AML cells from eight cases we demonstrate two types of GM-CSF receptors: one with low affinity (dissociation constant [kd] 5.1 to 24.8 nmol/L) and one with a high affinity (kd 31 to 104 pmol/L). These AML cells also expressed high affinity receptors for IL-3 (kd 24 to 104 pmol/L). Cross-competition experiments showed that an excess concentration of nonlabeled IL-3 completely prevented the high affinity binding of radiolabled GM-CSF. This competition occurred at 37 degrees C as well as 4 degrees C. Low affinity GM-CSF binding was not affected by IL-3. Binding of radiolabeled IL-3 could be prevented by nonlabeled GM-CSF. In certain cases, this competition was complete, whereas in others only partial (49% to 77%) reduction of the radiolabeled IL-3 binding was seen. On the basis of these ligand binding features, we propose the existence of three receptor types on AML cells: (1) low affinity GM-CSF receptors that do not bind IL-3, (2) dual high affinity GM-CSF/IL-3 receptors, and (3) high affinity IL-3 receptors that do not bind GM-CSF. We could also demonstrate these receptor types on normal monocytes. In addition, a fourth type of receptor was apparent on normal granulocytes (4), incapable of binding IL-3 and with an intermediate affinity for GM-CSF (approximately 400 pmol/L). Chemical crosslinking showed that GM-CSF and IL-3 both bind to proteins with molecular weight values of 130, 105, and 75, which provides additional evidence for the existence of a common GM-CSF/IL-3 receptor complex.
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PALMER, GENE C., JERRY A. MILLER, EDWARD F. CREGAN, PATRICIA GENDRON, and JAMES PEELING. "Low-Affinity NMDA Receptor Antagonists." Annals of the New York Academy of Sciences 825, no. 1 Neuroprotecti (October 1997): 220–31. http://dx.doi.org/10.1111/j.1749-6632.1997.tb48432.x.
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George, Andrew J. T., and David Gray. "Receptor editing during affinity maturation." Immunology Today 20, no. 4 (April 1999): 196. http://dx.doi.org/10.1016/s0167-5699(98)01408-x.
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Nachman, Michele, Abdul R. M. Azad, and Pascal Bailon. "Membrane-based receptor affinity chromatography." Journal of Chromatography A 597, no. 1-2 (April 1992): 155–66. http://dx.doi.org/10.1016/0021-9673(92)80105-4.
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Spence, Cheryl, Carol Ann Schaffer, Stephen Kessler, and Pascal Bailon. "Fluidized-bed receptor-affinity chromatography." Biomedical Chromatography 8, no. 5 (September 1994): 236–41. http://dx.doi.org/10.1002/bmc.1130080508.
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HANDA, RAJASH K. "Metabolism Alters the Selectivity of Angiotensin-(1-7) Receptor Ligands for Angiotensin Receptors." Journal of the American Society of Nephrology 11, no. 8 (August 2000): 1377–86. http://dx.doi.org/10.1681/asn.v1181377.
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Abstract. The present study examined whether metabolism of the putative angiotensin-(1-7) receptor agonist and antagonist [angiotensin-(1-7) and D-alanine7angiotensin-(1-7), respectively] altered their ability to interact with angiotensin AT1, AT2, and AT4receptor subtypes. Both angiotensin-(1-7) and D-alanine7angiotensin-(1-7) competed with low affinity for125I-sarcosine1, isoleucine8angiotensin II binding to AT1and AT2receptors in rat liver and adrenal medulla membranes, respectively, and competed with low affinity for125I-angiotensin IV binding to AT4receptors in bovine kidney epithelial cell membranes.In vitrorenal metabolism of the angiotensin-(1-7) receptor ligands (incubating peptides with rat cortical tissue homogenates) had minimal influence on low-affinity binding to AT1and AT2receptors, yet caused a significant and dramatic shift toward high-affinity binding for AT4receptors. Low-affinity angiotensin II binding to the AT4receptor was also shifted toward high-affinity binding following renal metabolism of the peptide. Conversely, angiotensins with high affinity for the AT4receptor (e.g., angiotensin IV) were shifted toward low-affinity binding states following peptide metabolism. Incubation of125I-angiotensin-(1-7) with rat cortical tissue generated the high-affinity AT4receptor ligand125I-angiotensin-(3-7), whereas the renal metabolism of125I-angiotensin II generated both125I-angiotensin-(3-7) and125I-angiotensin IV. These results reveal that renal metabolism of angiotensin-(1-7) receptor ligands and angiotensin II yields products that have high affinity for the AT4receptor and could potentially contribute to the biologic actions of the parent peptide in the kidney.
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Shen, Wen, and Malcolm M. Slaughter. "Internal Calcium Modulates Apparent Affinity of Metabotropic GABA Receptors." Journal of Neurophysiology 82, no. 6 (December 1, 1999): 3298–306. http://dx.doi.org/10.1152/jn.1999.82.6.3298.
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The metabotropic GABA receptor (GABABR) regulates calcium influx in neurons. Whole cell voltage-clamp techniques were employed to determine the effects of internal calcium on the activity of GABABRs. GABABR receptor apparent affinity was maximal when bis-( o-aminophenoxy)- N,N,N′,N′-tetraacetic acid (BAPTA) maintained internal calcium below 70 nM. Apparent affinity was reduced as internal calcium increased. EGTA did not produce similar effects, suggesting that localized increases in calcium influenced GABABR apparent affinity. Confocal imaging disclosed relatively high internal calcium just below the plasma membrane of isolated neurons. BAPTA, but not EGTA, reduced this ring of high calcium. Heparin, dantrolene, and ryanodine increased GABABR apparent affinity, effects similar to that of BAPTA. Calmodulin inhibitors also increased receptor apparent affinity. These results suggest that internally released calcium activates calmodulin, which reduces GABABR apparent affinity. This identifies a reciprocal system in which the metabotropic GABA receptor can reduce calcium influx, but internal calcium can suppress this receptor pathway. Metabotropic glutamate receptors linked to inositol 1,4,5 trisphosphate (InsP3) raised internal calcium and suppressed the action of GABABRs. Thus negative feedback systems control the balance between excitatory and inhibitory metabotropic receptor pathways in retinal neurons.
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Lee, Sangmin. "Development of High Affinity Calcitonin Analog Fragments Targeting Extracellular Domains of Calcitonin Family Receptors." Biomolecules 11, no. 9 (September 15, 2021): 1364. http://dx.doi.org/10.3390/biom11091364.
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The calcitonin and amylin receptors (CTR and AMY receptors) are the drug targets for osteoporosis and diabetes treatment, respectively. Salmon calcitonin (sCT) and pramlintide were developed as peptide drugs that activate these receptors. However, next-generation drugs with improved receptor binding profiles are desirable for more effective pharmacotherapy. The extracellular domain (ECD) of CTR was reported as the critical binding site for the C-terminal half of sCT. For the screening of high-affinity sCT analog fragments, purified CTR ECD was used for fluorescence polarization/anisotropy peptide binding assay. When three mutations (N26D, S29P, and P32HYP) were introduced to the sCT(22–32) fragment, sCT(22–32) affinity for the CTR ECD was increased by 21-fold. CTR was reported to form a complex with receptor activity-modifying protein (RAMP), and the CTR:RAMP complexes function as amylin receptors with increased binding for the peptide hormone amylin. All three types of functional AMY receptor ECDs were prepared and tested for the binding of the mutated sCT(22–32). Interestingly, the mutated sCT(22–32) also retained its high affinity for all three types of the AMY receptor ECDs. In summary, the mutated sCT(22–32) showing high affinity for CTR and AMY receptor ECDs could be considered for developing the next-generation peptide agonists.
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Yu, D. H., S. C. Huang, S. A. Wank, S. Mantey, J. D. Gardner, and R. T. Jensen. "Pancreatic receptors for cholecystokinin: evidence for three receptor classes." American Journal of Physiology-Gastrointestinal and Liver Physiology 258, no. 1 (January 1, 1990): G86—G95. http://dx.doi.org/10.1152/ajpgi.1990.258.1.g86.
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For inhibition of binding of 125I-Bolton-Hunter-labeled cholecystokinin octapeptide (125I-BH-CCK-8) to guinea pig pancreatic acini, the potencies for agonists were CCK-8 greater than desulfated [des(SO3)] CCK-8 greater than gastrin-17-I greater than pentagastrin greater than CCK-4 and for the antagonists L 364718 greater than proglumide analogue 10 greater than CBZ-CCK-(27-32)-NH2. For all non-sulfated agonists, the curves were biphasic with 20% of the tracer bound to sites with high affinity for these agonists with the following relative potencies: gastrin-17-I greater than pentagastrin greater than des(SO3)CCK-8 much greater than CCK-4; whereas 80% was bound to low-affinity sites with the following potencies: des(SO3)CCK-8 greater than gastrin-17-I = pentagastrin much greater than CCK-4. For L 364718 and proglumide analogue 10, 80% of 125I-BH-CCK-8 was bound to sites with high affinity for these antagonists and 20% to sites with low affinity. Analysis of the dose-inhibition curve for CCK-8 demonstrated two binding sites; however, comparison with the analysis in the presence of 0.1 microM gastrin-17-I suggested three binding sites. The gastrin-17-I dose-inhibition curve was significantly better fit by a three-site model than by a two-site model. The affinities of the various agonists and antagonists for the three sites were compared with their abilities to inhibit binding of 125I-gastrin-I and either stimulate or inhibit CCK-8-stimulated amylase release. These results demonstrate that 125I-BH-CCK-8 binds to three classes of receptors, not two as reported previously. Two classes are CCK-preferring, bind 83% of 125I-BH-CCK-8 at tracer concentrations, and comprise high- and low-affinity CCK-preferring sites that can be distinguished by all agonists but have equally high affinity for L 364718 and proglumide 10. A third class binds 17% of the tracer, cannot be differentiated from high-affinity CCK-preferring receptors by CCK-8, and has low affinities for L 364718 and proglumide 10. Future studies relating binding of 125I-BH-CCK-8 to biological activity or characterization of the CCK receptor by using radiolabeled agonists should consider CCK interaction with three receptors, not two as was done in the past.
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Montiel, F., J. Ortiz-Caro, A. Villa, A. Pascual, and A. Aranda. "Presence of insulin receptors in cultured glial C6 cells. Regulation by butyrate." Biochemical Journal 258, no. 1 (February 15, 1989): 147–55. http://dx.doi.org/10.1042/bj2580147.
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The presence of insulin receptor and its regulation by butyrate and other short-chain fatty acids was studied in C6 cells, a rat glioma cell line. Intact C6 cells bind 125I-insulin in a rapid, reversible and specific manner. Scatchard analysis of the binding data gives typical curvilinear plots with apparent affinities of approx. 6 nM and 70 nM for the low-affinity (approx. 90% of total) and high-affinity (approx. 10% of total) sites respectively. Incubation with butyrate results in a time- and dose-dependent decrease of insulin binding to C6 cells. A maximal effect was found with 2 mM-butyrate that decreased the receptor by 40-70% after 48 h. Butyrate decreased numbers of receptors of both classes, but did not significantly alter receptor affinity. Other short-chain fatty acids, as well as keto acids, had a similar effect, but with a lower potency. Cycloheximide caused an accumulation of insulin receptors at the cell surface, since insulin binding increased and receptor affinity did not change after incubation with the inhibitor. Simultaneous addition of butyrate and cycloheximide abolished the loss of receptors produced by the fatty acid. In cells preincubated with butyrate, cycloheximide also produced a large increase in receptor numbers, showing that in the absence of new receptor synthesis a large pool of receptors re-appears at the surface of butyrate-treated cells.
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Nagler, A., L. L. Lanier, and J. H. Phillips. "Constitutive expression of high affinity interleukin 2 receptors on human CD16-natural killer cells in vivo." Journal of Experimental Medicine 171, no. 5 (May 1, 1990): 1527–33. http://dx.doi.org/10.1084/jem.171.5.1527.
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The majority of human NK cells express low affinity IgG Fc receptors (CD16+), whereas a minor subset of NK cells lack Fc receptor expression (CD16-). In contrast to CD16+ NK cells that express only p75 IL-2 receptors, CD16- NK cells constitutively co-express both p75 and p55 IL-2 receptors in vivo and preferentially respond to low concentrations of IL-2 with increased cytolytic activation and proliferation. Scatchard analysis demonstrated the presence of approximately 1,200 high affinity (approximately 25 pM kD) and approximately 9,600 intermediate affinity (approximately 2 nM kD) IL-2 receptors on CD16- NK cells. CD16+ NK cells expressed only a single intermediate affinity IL-2 receptor of approximately 1.9 nM kD (approximately 9,000 sites per cell). The IL-2 binding data thus substantiated the phenotypic and functional studies and definitively show that the differential responsiveness of CD16- and CD16+ NK cells to IL-2 is manifested through different affinity IL-2 receptors.
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Antoni, F. A., and S. E. Chadio. "Essential role of magnesium in oxytocin-receptor affinity and ligand specificity." Biochemical Journal 257, no. 2 (January 15, 1989): 611–14. http://dx.doi.org/10.1042/bj2570611.
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We have analysed, with the aid of a novel radioiodinated oxytocin (OT)-receptor antagonist, the role of Mg2+ in uterine OT-receptor function. The antagonist-receptor interaction was characterized by high affinity, reversibility and stereospecificity in Tris/HCl buffer containing 3 mmol of Mg2+/litre as well as buffer free of Mg2+. By contrast, omission of Mg2+ decreased the affinity of the receptor for OT by about 1500-fold; moreover, the stereospecificity of agonist, but not antagonist, binding was lost. Since guanine nucleotides had relatively minor effects in this system (less than or equal to 2-fold decrease in OT affinity), we suggest that the agonist-binding site of OT receptors is directly modulated by Mg2+, unlike other receptors, where the effects of bivalent cations are exerted via guanine-nucleotide-binding (G-) proteins. Thus the ligand recognition mechanism of OT receptors may be novel in this respect.
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Lu, Jessica, Sarah J. Piper, Peishen Zhao, Laurence J. Miller, Denise Wootten, and Patrick M. Sexton. "Targeting VIP and PACAP Receptor Signaling: New Insights into Designing Drugs for the PACAP Subfamily of Receptors." International Journal of Molecular Sciences 23, no. 15 (July 22, 2022): 8069. http://dx.doi.org/10.3390/ijms23158069.
Full textAbstract:
Pituitary Adenylate Cyclase-Activating Peptide (PACAP) and Vasoactive Intestinal Peptide (VIP) are neuropeptides involved in a diverse array of physiological and pathological processes through activating the PACAP subfamily of class B1 G protein-coupled receptors (GPCRs): VIP receptor 1 (VPAC1R), VIP receptor 2 (VPAC2R), and PACAP type I receptor (PAC1R). VIP and PACAP share nearly 70% amino acid sequence identity, while their receptors PAC1R, VPAC1R, and VPAC2R share 60% homology in the transmembrane regions of the receptor. PACAP binds with high affinity to all three receptors, while VIP binds with high affinity to VPAC1R and VPAC2R, and has a thousand-fold lower affinity for PAC1R compared to PACAP. Due to the wide distribution of VIP and PACAP receptors in the body, potential therapeutic applications of drugs targeting these receptors, as well as expected undesired side effects, are numerous. Designing selective therapeutics targeting these receptors remains challenging due to their structural similarities. This review discusses recent discoveries on the molecular mechanisms involved in the selectivity and signaling of the PACAP subfamily of receptors, and future considerations for therapeutic targeting.
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Kitsukawa, Y., Z. F. Gu, P. Hildebrand, and R. T. Jensen. "Gastric smooth muscle cells possess two classes of endothelin receptors but only one alters contraction." American Journal of Physiology-Gastrointestinal and Liver Physiology 266, no. 4 (April 1, 1994): G713—G721. http://dx.doi.org/10.1152/ajpgi.1994.266.4.g713.
Full textAbstract:
Endothelin (ET)-like immunoreactivity and ET binding sites are widely distributed in the gastrointestinal tract, and ET causes contraction of stomach muscle strips. To determine whether ETs could interact with gastric smooth muscle cells directly and alter function, we measured binding of 125I-ET-1, 125I-ET-2, and 125I-ET-3 to dispersed gastric smooth muscle cells from guinea pig and their abilities to alter cell length. Each ligand bound in a time- and temperature-dependent manner, which was specific and saturable. Analysis of the dose-inhibition curves of both ET-1 and ET-3 for binding of each ligand indicated the presence of two classes of receptors, one class (ETA receptor) with a high affinity for ET-1 and ET-2 but a low affinity for ET-3, and the other (ETB receptor) with a high affinity for ET-1, ET-2, and ET-3. The ligands were rapidly internalized by both receptors; however, it was greater with ETA receptors. ET-1 stimulated muscle contraction (50% effective concentration approximately 2 nM), whereas ET-3 did not stimulate contraction or cause relaxation. These results demonstrate that gastric smooth muscle cells possess two classes of ET receptors. One type (ETA) has a high affinity for ET-1 and ET-2 and a low affinity for ET-3, and receptor occupation results in rapid ligand internalization and muscle contraction; the other type (ETB) has a high affinity for ET-1, ET-2, and ET-3, and receptor occupation results in a lesser degree of ligand internalization than the ETA receptor and does not alter contractile behavior.
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McGrattan, P. D., and A. R. G. Wylie. "Influence of genotype and gender on receptor binding affinity (Kd) for insulin and insulin receptor concentration in skeletal muscle and adipose tissue of growing lambs." Animal Science 73, no. 1 (April 2001): 77–84. http://dx.doi.org/10.1017/s1357729800058070.
Full textAbstract:
AbstractDifferences in receptor binding affinity for insulin, and insulin receptor concentration, were found in skeletal muscle and adipose tissue of lambs differing in genotype and gender. The Rouge de l’Ouest lamb genotype had a lower receptor affinity for insulin in m. rectus capitis skeletal muscle compared with both Dutch Texel and Greyface lamb genotypes (P < 0·01). Insulin receptor concentration in m. longissimus dorsi skeletal muscle was greater in the Texel lamb genotype compared with both the Greyface (P < 0·001) and Rouge de l’Ouest (P < 0·05) lamb genotypes. Insulin receptor concentration in perirenal fat tissue was greater in the Rouge de l’Ouest lamb genotype compared with the Texel lamb genotype (P < 0·05).There were no differences in receptor affinity for insulin between individual tissues in male and female lambs. The concentration of insulin receptors in male lambs was greater in m. longissimus dorsi (P < 0·05), subcutaneous adipose tissue (P < 0·05) and perirenal fat (P < 0·001) compared with female lambs. Receptor affinity for insulin in individual tissue depots also demonstrated significant genotype ✕ gender interactions. M. longissimus dorsi had a lower receptor affinity for insulin in Texel male lambs compared with Greyface male lambs (P < 0·05). M. rectus capitis had a lower receptor affinity in Rouge de l’Ouest male lambs compared with Texel male lambs (P < 0·01). M. rectus capitis also had a lower receptor affinity for insulin in Rouge de l’Ouest female lambs compared with both Texel and Greyface female lambs (P < 0·01). Within individual genotypes, m. longissimus dorsi had a lower receptor affinity for insulin in Greyface female lambs compared with Greyface male lambs (P < 0·05) whereas m. rectus capitis had a lower receptor affinity for insulin in Texel female lambs compared with Texel male lambs (P < 0·05).Such differences in tissue receptor affinity for insulin and receptor concentration in lambs differing in genotype and gender could, through effects on tissue sensitivity and/or responsiveness to insulin, manipulate nutrient partitioning between skeletal muscle and fat tissue and hence control, at least in part, the efficiency of lean meat production.
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Faull, R. J., N. L. Kovach, J. M. Harlan, and M. H. Ginsberg. "Affinity modulation of integrin alpha 5 beta 1: regulation of the functional response by soluble fibronectin." Journal of Cell Biology 121, no. 1 (April 1, 1993): 155–62. http://dx.doi.org/10.1083/jcb.121.1.155.
Full textAbstract:
We report that a beta 1 integrin (alpha 5 beta 1) can exist in different affinity states for its soluble ligand, fibronectin. The alpha 5 beta 1 expressed by the erythroleukemic cell line K562 binds soluble fibronectin with low affinity (Kd > 1 microM), but is induced to bind it with 20-fold higher affinity (Kd-54 nM) in the presence of the anti-beta 1 mAb 8A2. This activation seems to be due to direct antibody-induced change in the receptor that does not require intracellular signaling, and is a plausible basis for the 8A2-induced enhancement of beta 1-dependent adhesion to fibronectin and other immobilized ligands (Kovach, N. L., T. M. Carlos, E. Yee, and J. M. Harlan. 1992. J. Cell Biol. 116: 499-509). Fab fragments of 8A2 bind with higher affinity to alpha 5 beta 1 receptor that is occupied by the GRG-DSP peptide ligand suggesting that the antibody functions by stabilizing a high affinity (occupied) conformer of the receptor. A functional consequence of the affinity modulation is that soluble fibronectin (at physiological concentrations) occupies the high affinity receptors, and so becomes an effective inhibitor of adhesion to immobilized fibronectin. In contrast, the majority of low affinity receptors remain unoccupied and are still to mediate cellular adhesion.
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Spivak, JL, LS Avedissian, JH Pierce, D. Williams, WD Hankins, and RA Jensen. "Isolation of the full-length murine erythropoietin receptor using a baculovirus expression system." Blood 87, no. 3 (February 1, 1996): 926–37. http://dx.doi.org/10.1182/blood.v87.3.926.bloodjournal873926.
Full textAbstract:
The full-length murine erythropoietin receptor was expressed in Spodoptera frugiperda (Sf9) cells using a recombinant baculovirus vector. Erythropoietin receptor protein production was maximal 48 hours after infection, as determined by metabolic labeling and immunoblotting; receptor protein varied in molecular mass from 62 to 76 kD. Erythropoietin receptors produced in Sf9 cells could be solubilized using CHAPS in a form capable of binding erythropoietin, and the solubilized receptor bound to immobilized Concanavalin A (Con A) and wheat germ agglutinin, as well as to immobilized recombinant human erythropoietin. Analysis of the distribution of erythropoietin receptors in Sf9 plasma membrane and cytosol fractions using lectin affinity chromatography revealed that membrane-bound receptor had a higher apparent molecular mass and contained the bulk of receptors that bound to wheat germ agglutinin. The receptor was purified by sequential affinity chromatography on Con A-Sepharose and immobilized erythropoietin. Erythropoietin receptors expressed in Sf9 cells were inserted into the plasma membrane in the correct orientation, bound 125I-erythropoietin with a single affinity (kD, 330 pmol/L), and were internalized after ligand binding. However, kD varied inversely with the number of cell surface receptors. Solubilized erythropoietin receptors in whole-cell lysates and isolated plasma membranes exhibited high-affinity binding, with kD values of 92 and 57 pmol/L, respectively. Erythropoietin bound to the surface of infected Sf9 cells could be cross-linked to two proteins with molecular masses of 90 and 65 kD using the homobifunctional cross-linker, disuccinimidyl suberate (DSS). Similar results were obtained with solubilized receptors in whole-cell lysates, and both proteins could be immunoprecipitated by an antiserum to the erythropoietin receptor carboxyl-terminal domain.