Allosteric interactions between the binding sites of receptor agonists and guanine nucleotides: A comparative study of the 5-hydroxytryptamine(1A) and adenosine A(1) receptor systems.
Item
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Title
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Allosteric interactions between the binding sites of receptor agonists and guanine nucleotides: A comparative study of the 5-hydroxytryptamine(1A) and adenosine A(1) receptor systems.
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Identifier
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AAI9207100
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identifier
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9207100
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Creator
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Mahle, Cathy Diane.
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Contributor
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Adviser: Saul Maayani
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Date
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1991
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Language
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English
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Publisher
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City University of New York.
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Subject
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Health Sciences, Pharmacology
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Abstract
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Receptor-mediated signal transduction requires interaction between receptor, G-protein, and effector. Agonist binding to receptor stabilizes a high affinity ternary complex, H-R-G. Guanine nucleotides (GN) destablize H-R-G, converting the high affinity into the low affinity state. Characteristics of the allosteric interaction between binding of hormone to receptor and GN to G-protein(s) were investigated.;Initially this interaction was examined in rat hippocampal membranes, using three ({dollar}\sp3{dollar}H) -agonists: an AD A{dollar}\sb1{dollar} agonist ( ({dollar}\sp3{dollar}H) -R-PIA), and a full ( ({dollar}\sp3{dollar}H) -8-OH-DPAT) and partial ( ({dollar}\sp3{dollar}H) -BMY-7378) 5-HT{dollar}\sb{lcub}\rm 1A{rcub}{dollar} agonist. Density (B{dollar}\sb{lcub}\rm max{rcub}{dollar}, pmol/mg protein) of H-R-G appeared dependent on receptor type and agonist relative efficacy. GN decreased H-R-G density (50%), while K{dollar}\sb{lcub}\rm d{rcub}{dollar} values remained constant.;GN attenuated ({dollar}\sp3{dollar}H) -agonist binding in a concentration-dependent saturable manner. Parameters of the attenuation (IC{dollar}\sb{lcub}50{rcub}{dollar}, E{dollar}\sb{lcub}\rm max{rcub}{dollar} and slope index) were used to characterize the allosterism occurring between binding of agonist to receptor, and GN to G-protein(s). Similar IC{dollar}\sb{lcub}50{rcub}{dollar} values were observed for 5-HT{dollar}\sb{lcub}\rm 1A{rcub}{dollar} agonists. Higher IC{dollar}\sb{lcub}50{rcub}{dollar} values were observed with the AD A{dollar}\sb1{dollar} agonist at equivalent receptor occupancy. IC{dollar}\sb{lcub}50{rcub}{dollar} values increased with increasing receptor occupancy; IC{dollar}\sb{lcub}50{rcub}{dollar} ({dollar}\mu{dollar}M): R-PIA 0.6-2.6; 8-OH-DPAT, BMY-7378 0.18-0.8, indicating IC{dollar}\sb{lcub}50{rcub}{dollar} values were effected by degree of agonist occupancy, receptor type, and not drug relative efficacy. GTP-dependent inhibition of adenylyl cyclase was indistinguishable across agonist (EC{dollar}\sb{lcub}50{rcub}{dollar} 0.3-0.5 {dollar}\mu{dollar}M). Events related to drug relative efficacy may occur at R/G interaction and not GN/G-protein or G-protein/effector.;Quantitative receptor autoradiography (QRA) was applied to study this allosterism in rat and human hippocampal subregions. Agonist affinity was similar in hippocampal membranes and subregions. Parameters of GN concentration-effect curves were determined. In contrast to membrane preparations, attenuation of ({dollar}\sp3{dollar}H) -agonist binding by Gpp(NH)p in hippocampal subregions was nearly complete. Within regions, IC{dollar}\sb{lcub}50{rcub}{dollar} values were indistinguishable, but for ({dollar}\sp3{dollar}H) -8-OH-DPAT were lower (0.1-0.2 {dollar}\mu{dollar}M) in human than rat (0.3-0.4 {dollar}\mu{dollar}M). In human hippocampus the IC{dollar}\sb{lcub}50{rcub}{dollar} of the GN curves was lower for ({dollar}\sp3{dollar}H) -8-OH-DPAT than ({dollar}\sp3{dollar}H) -R-PIA (0.2-0.3 {dollar}\mu{dollar}M). Allosteric interactions between binding of hormone and GN to the receptor system can be measured using QRA, enabling discrete localization and characterization of H-R-G.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
