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Advances in Methods to Characterize Ligand-Induced Ionic Lock and Rotamer Toggle Molecular Switch in G Protein-Coupled Receptors

Structural biology of GPCRs has made significant progress upon recently developed technologies for GPCRs expression/purification and elucidation of GPCRs crystal structures. The crystal structures provide a snapshot of the receptor structural disposition of GPCRs itself or with cocrystallized ligand... Full description

Journal Title: Methods in Enzymology 2013, Vol.520, p.153-174
Main Author: Xie, Xiang-Qun
Other Authors: Chowdhury, Ananda
Format: Electronic Article Electronic Article
Language: English
Subjects:
sf9
Quelle: Gale eBooks
Publisher: United States: Elsevier Science & Technology
ID: ISSN: 0076-6879
Link: https://www.ncbi.nlm.nih.gov/pubmed/23332699
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recordid: cdi_gale_vrl_7002400016
title: Advances in Methods to Characterize Ligand-Induced Ionic Lock and Rotamer Toggle Molecular Switch in G Protein-Coupled Receptors
format: Article
creator:
  • Xie, Xiang-Qun
  • Chowdhury, Ananda
subjects:
  • Animals
  • Article
  • expression
  • G Protein Coupled Receptors (GPCR)
  • G proteins
  • hormone antagonists
  • hormone substitutes
  • hormones
  • Humans
  • Ionic Lock
  • Molecular Switches
  • Protein Binding - genetics
  • Protein Binding - physiology
  • Protein Structure, Secondary
  • purifications
  • Receptors, Adrenergic, beta-2 - chemistry
  • Receptors, Adrenergic, beta-2 - genetics
  • Receptors, Adrenergic, beta-2 - metabolism
  • Receptors, G-Protein-Coupled - chemistry
  • Receptors, G-Protein-Coupled - genetics
  • Receptors, G-Protein-Coupled - metabolism
  • recombinant membrane protein
  • Rotamer Toggle
  • sf9
  • Signal Transduction - genetics
  • Signal Transduction - physiology
  • Switches
ispartof: Methods in Enzymology, 2013, Vol.520, p.153-174
description: Structural biology of GPCRs has made significant progress upon recently developed technologies for GPCRs expression/purification and elucidation of GPCRs crystal structures. The crystal structures provide a snapshot of the receptor structural disposition of GPCRs itself or with cocrystallized ligands, and the results are congruent with biophysical and computer modeling studies reported about GPCRs conformational and dynamics flexibility, regulated activation, and the various stabilizing interactions, such as “molecular switches.” The molecular switches generally constitute the most conserved domains within a particular GPCR superfamily. Often agonist-induced receptor activation proceeds by the disruption of majority of these interactions, while antagonist and inverse agonist act as blockers and structural stabilizers, respectively. Several elegant studies, particularly for the β2AR, have demonstrated the relationship between ligand structure, receptor conformational changes, and corresponding pharmacological outcomes. Thus, it is of great importance to understand GPCRs activation related to cell signaling pathways. Herein, we summarize the steps to produce functional GPCRs, generate suitably fluorescent labeled GPCRs and the procedure to use that to understand if ligand-induced activation can proceed by activation of the GPCRs via ionic lock switch and/or rotamer toggle switch mechanisms. Such understanding of ligand structure and mechanism of receptor activation will provide great insight toward uncovering newer pathways of GPCR activation and aid in structure-based drug design.
language: eng
source: Gale eBooks
identifier: ISSN: 0076-6879
fulltext: fulltext
issn:
  • 0076-6879
  • 1557-7988
url: Link


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descriptionStructural biology of GPCRs has made significant progress upon recently developed technologies for GPCRs expression/purification and elucidation of GPCRs crystal structures. The crystal structures provide a snapshot of the receptor structural disposition of GPCRs itself or with cocrystallized ligands, and the results are congruent with biophysical and computer modeling studies reported about GPCRs conformational and dynamics flexibility, regulated activation, and the various stabilizing interactions, such as “molecular switches.” The molecular switches generally constitute the most conserved domains within a particular GPCR superfamily. Often agonist-induced receptor activation proceeds by the disruption of majority of these interactions, while antagonist and inverse agonist act as blockers and structural stabilizers, respectively. Several elegant studies, particularly for the β2AR, have demonstrated the relationship between ligand structure, receptor conformational changes, and corresponding pharmacological outcomes. Thus, it is of great importance to understand GPCRs activation related to cell signaling pathways. Herein, we summarize the steps to produce functional GPCRs, generate suitably fluorescent labeled GPCRs and the procedure to use that to understand if ligand-induced activation can proceed by activation of the GPCRs via ionic lock switch and/or rotamer toggle switch mechanisms. Such understanding of ligand structure and mechanism of receptor activation will provide great insight toward uncovering newer pathways of GPCR activation and aid in structure-based drug design.
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languageeng
publisherUnited States: Elsevier Science & Technology
subjectAnimals ; Article ; expression ; G Protein Coupled Receptors (GPCR) ; G proteins ; hormone antagonists ; hormone substitutes ; hormones ; Humans ; Ionic Lock ; Molecular Switches ; Protein Binding - genetics ; Protein Binding - physiology ; Protein Structure, Secondary ; purifications ; Receptors, Adrenergic, beta-2 - chemistry ; Receptors, Adrenergic, beta-2 - genetics ; Receptors, Adrenergic, beta-2 - metabolism ; Receptors, G-Protein-Coupled - chemistry ; Receptors, G-Protein-Coupled - genetics ; Receptors, G-Protein-Coupled - metabolism ; recombinant membrane protein ; Rotamer Toggle ; sf9 ; Signal Transduction - genetics ; Signal Transduction - physiology ; Switches
ispartofMethods in Enzymology, 2013, Vol.520, p.153-174
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descriptionStructural biology of GPCRs has made significant progress upon recently developed technologies for GPCRs expression/purification and elucidation of GPCRs crystal structures. The crystal structures provide a snapshot of the receptor structural disposition of GPCRs itself or with cocrystallized ligands, and the results are congruent with biophysical and computer modeling studies reported about GPCRs conformational and dynamics flexibility, regulated activation, and the various stabilizing interactions, such as “molecular switches.” The molecular switches generally constitute the most conserved domains within a particular GPCR superfamily. Often agonist-induced receptor activation proceeds by the disruption of majority of these interactions, while antagonist and inverse agonist act as blockers and structural stabilizers, respectively. Several elegant studies, particularly for the β2AR, have demonstrated the relationship between ligand structure, receptor conformational changes, and corresponding pharmacological outcomes. Thus, it is of great importance to understand GPCRs activation related to cell signaling pathways. Herein, we summarize the steps to produce functional GPCRs, generate suitably fluorescent labeled GPCRs and the procedure to use that to understand if ligand-induced activation can proceed by activation of the GPCRs via ionic lock switch and/or rotamer toggle switch mechanisms. Such understanding of ligand structure and mechanism of receptor activation will provide great insight toward uncovering newer pathways of GPCR activation and aid in structure-based drug design.
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abstractStructural biology of GPCRs has made significant progress upon recently developed technologies for GPCRs expression/purification and elucidation of GPCRs crystal structures. The crystal structures provide a snapshot of the receptor structural disposition of GPCRs itself or with cocrystallized ligands, and the results are congruent with biophysical and computer modeling studies reported about GPCRs conformational and dynamics flexibility, regulated activation, and the various stabilizing interactions, such as “molecular switches.” The molecular switches generally constitute the most conserved domains within a particular GPCR superfamily. Often agonist-induced receptor activation proceeds by the disruption of majority of these interactions, while antagonist and inverse agonist act as blockers and structural stabilizers, respectively. Several elegant studies, particularly for the β2AR, have demonstrated the relationship between ligand structure, receptor conformational changes, and corresponding pharmacological outcomes. Thus, it is of great importance to understand GPCRs activation related to cell signaling pathways. Herein, we summarize the steps to produce functional GPCRs, generate suitably fluorescent labeled GPCRs and the procedure to use that to understand if ligand-induced activation can proceed by activation of the GPCRs via ionic lock switch and/or rotamer toggle switch mechanisms. Such understanding of ligand structure and mechanism of receptor activation will provide great insight toward uncovering newer pathways of GPCR activation and aid in structure-based drug design.
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