Viewing rare conformations of the β 2 adrenergic receptor with pressure-resolved DEER spectroscopy.

Autor: Lerch MT; Jules Stein Eye Institute, University of California, Los Angeles, CA 90095.; Department of Chemistry and Biochemistry, University of California, Los Angeles, CA 90095., Matt RA; Department of Chemical and Systems Biology, Stanford University School of Medicine, Stanford, CA 94305., Masureel M; Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305., Elgeti M; Jules Stein Eye Institute, University of California, Los Angeles, CA 90095.; Department of Chemistry and Biochemistry, University of California, Los Angeles, CA 90095., Kumar KK; Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305., Hilger D; Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305., Foys B; Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305., Kobilka BK; Department of Chemical and Systems Biology, Stanford University School of Medicine, Stanford, CA 94305; kobilka@stanford.edu hubbellw@jsei.ucla.edu.; Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305., Hubbell WL; Jules Stein Eye Institute, University of California, Los Angeles, CA 90095; kobilka@stanford.edu hubbellw@jsei.ucla.edu.; Department of Chemistry and Biochemistry, University of California, Los Angeles, CA 90095.
Jazyk: angličtina
Zdroj: Proceedings of the National Academy of Sciences of the United States of America [Proc Natl Acad Sci U S A] 2020 Dec 15; Vol. 117 (50), pp. 31824-31831. Date of Electronic Publication: 2020 Nov 30.
DOI: 10.1073/pnas.2013904117
Abstrakt: The β 2 adrenergic receptor (β 2 AR) is an archetypal G protein coupled receptor (GPCR). One structural signature of GPCR activation is a large-scale movement (ca. 6 to 14 Å) of transmembrane helix 6 (TM6) to a conformation which binds and activates a cognate G protein. The β 2 AR exhibits a low level of agonist-independent G protein activation. The structural origin of this basal activity and its suppression by inverse agonists is unknown but could involve a unique receptor conformation that promotes G protein activation. Alternatively, a conformational selection model proposes that a minor population of the canonical active receptor conformation exists in equilibrium with inactive forms, thus giving rise to basal activity of the ligand-free receptor. Previous spin-labeling and fluorescence resonance energy transfer experiments designed to monitor the positional distribution of TM6 did not detect the presence of the active conformation of ligand-free β 2 AR. Here we employ spin-labeling and pressure-resolved double electron-electron resonance spectroscopy to reveal the presence of a minor population of unliganded receptor, with the signature outward TM6 displacement, in equilibrium with inactive conformations. Binding of inverse agonists suppresses this population. These results provide direct structural evidence in favor of a conformational selection model for basal activity in β 2 AR and provide a mechanism for inverse agonism. In addition, they emphasize 1) the importance of minor populations in GPCR catalytic function; 2) the use of spin-labeling and variable-pressure electron paramagnetic resonance to reveal them in a membrane protein; and 3) the quantitative evaluation of their thermodynamic properties relative to the inactive forms, including free energy, partial molar volume, and compressibility.
Competing Interests: Competing interest statement: B.K.K. is a cofounder of and consultant for ConfometRx, Inc.
Databáze: MEDLINE
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