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Nat Methods. 2015 Sep;12(9):845-851. doi: 10.1038/nmeth.3479. Epub 2015 Jul 13.

Imaging G protein-coupled receptors while quantifying their ligand-binding free-energy landscape.

Author information

1
Department of Biosystems Science and Engineering, Eidgenössische Technische Hochschule (ETH) Zurich, Basel, Switzerland.
2
Department of Cellular Physiology and Medicine, Stanford University School of Medicine, Stanford, California, USA.
3
Cardiovascular Research Institute, University of California, San Francisco, San Francisco, California, USA.
#
Contributed equally

Abstract

Imaging native membrane receptors and testing how they interact with ligands is of fundamental interest in the life sciences but has proven remarkably difficult to accomplish. Here, we introduce an approach that uses force-distance curve-based atomic force microscopy to simultaneously image single native G protein-coupled receptors in membranes and quantify their dynamic binding strength to native and synthetic ligands. We measured kinetic and thermodynamic parameters for individual protease-activated receptor-1 (PAR1) molecules in the absence and presence of antagonists, and these measurements enabled us to describe PAR1's ligand-binding free-energy landscape with high accuracy. Our nanoscopic method opens an avenue to directly image and characterize ligand binding of native membrane receptors.

PMID:
26167642
PMCID:
PMC5087271
DOI:
10.1038/nmeth.3479
[Indexed for MEDLINE]
Free PMC Article

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