Publication Date:
2011-01-14
Description:
G-protein-coupled receptors (GPCRs) are eukaryotic integral membrane proteins that modulate biological function by initiating cellular signalling in response to chemically diverse agonists. Despite recent progress in the structural biology of GPCRs, the molecular basis for agonist binding and allosteric modulation of these proteins is poorly understood. Structural knowledge of agonist-bound states is essential for deciphering the mechanism of receptor activation, and for structure-guided design and optimization of ligands. However, the crystallization of agonist-bound GPCRs has been hampered by modest affinities and rapid off-rates of available agonists. Using the inactive structure of the human beta(2) adrenergic receptor (beta(2)AR) as a guide, we designed a beta(2)AR agonist that can be covalently tethered to a specific site on the receptor through a disulphide bond. The covalent beta(2)AR-agonist complex forms efficiently, and is capable of activating a heterotrimeric G protein. We crystallized a covalent agonist-bound beta(2)AR-T4L fusion protein in lipid bilayers through the use of the lipidic mesophase method, and determined its structure at 3.5 A resolution. A comparison to the inactive structure and an antibody-stabilized active structure (companion paper) shows how binding events at both the extracellular and intracellular surfaces are required to stabilize an active conformation of the receptor. The structures are in agreement with long-timescale (up to 30 mus) molecular dynamics simulations showing that an agonist-bound active conformation spontaneously relaxes to an inactive-like conformation in the absence of a G protein or stabilizing antibody.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3074335/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉 〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3074335/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Rosenbaum, Daniel M -- Zhang, Cheng -- Lyons, Joseph A -- Holl, Ralph -- Aragao, David -- Arlow, Daniel H -- Rasmussen, Soren G F -- Choi, Hee-Jung -- Devree, Brian T -- Sunahara, Roger K -- Chae, Pil Seok -- Gellman, Samuel H -- Dror, Ron O -- Shaw, David E -- Weis, William I -- Caffrey, Martin -- Gmeiner, Peter -- Kobilka, Brian K -- 50GM073210/GM/NIGMS NIH HHS/ -- GM56169/GM/NIGMS NIH HHS/ -- GM75915/GM/NIGMS NIH HHS/ -- M083118/PHS HHS/ -- NS028471/NS/NINDS NIH HHS/ -- P01 GM75913/GM/NIGMS NIH HHS/ -- P60DK-20572/DK/NIDDK NIH HHS/ -- R01 GM068603/GM/NIGMS NIH HHS/ -- R37 NS028471/NS/NINDS NIH HHS/ -- R37 NS028471-20/NS/NINDS NIH HHS/ -- England -- Nature. 2011 Jan 13;469(7329):236-40. doi: 10.1038/nature09665.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Molecular and Cellular Physiology, Stanford University School of Medicine, 279 Campus Drive, Stanford, California 94305, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/21228876" target="_blank"〉PubMed〈/a〉
Keywords:
Adrenergic beta-2 Receptor Agonists/*chemistry/*metabolism
;
Crystallization
;
Crystallography, X-Ray
;
Disulfides/chemistry/metabolism
;
Drug Inverse Agonism
;
Heterotrimeric GTP-Binding Proteins/metabolism
;
Humans
;
Lipid Bilayers/chemistry/metabolism
;
Models, Molecular
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Molecular Dynamics Simulation
;
Procaterol/chemistry/metabolism
;
Propanolamines/chemistry/metabolism
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Protein Conformation
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Receptors, Adrenergic, beta-2/*chemistry/*metabolism
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Recombinant Fusion Proteins/chemistry/metabolism
;
Viral Proteins/chemistry/metabolism
Print ISSN:
0028-0836
Electronic ISSN:
1476-4687
Topics:
Biology
,
Chemistry and Pharmacology
,
Medicine
,
Natural Sciences in General
,
Physics
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