Single-molecule force spectroscopy study of interactions between angiotensin II type 1 receptor and different biased ligands in living cells.
Anal Bioanal Chem
; 410(14): 3275-3284, 2018 May.
Article
em En
| MEDLINE
| ID: mdl-29492619
ABSTRACT
Angiotensin II type 1 receptor (AT1R), a typical G protein-coupled receptor, plays a key role in regulating many cardiovascular functions. Different ligands can bind with AT1R to selectively activate either G protein (Gq) or ß-arrestin (ß-arr) pathway, or both pathways, but the molecular mechanism is not clear yet. In this work, we used, for the first time, atomic force microscopy-based single molecule force spectroscopy (SMFS) to study the interactions of AT1R with three types of ligands, balanced ligand, Gq-biased ligand, and ß-arr-biased ligand, in living cells. The results revealed their difference in binding force and binding stability. The complex of the Gq-biased ligand-AT1R overcame two energy barriers with an intermediate state during dissociation, whereas that of ß-arr-biased ligand-AT1R complex overcame one energy barrier. This indicated that AT1R had different ligand-binding conformational substates and underwent different structural changes to activate downstream signaling pathways with variable agonist efficacies. Quantitative analysis of AT1R-ligand binding in living cells at the single-molecule level offers a new tool to study the molecular mechanism of AT1R biased activation. Graphical Abstract Single-molecule force measurement on the living cell expressing AT1R-eGFP with a ligand modified AFM tip (left), the dynamic force spectra of ß-arrestin biased ligands-AT1R (middle), and Gq-biased ligands-AT1R (right). The complexes of ß-arr-biased ligand-AT1R overcame one energy barrier, with one linear region in the spectra, whereas the Gq-biased ligand-AT1R complexes overcame two energy barriers with two linear regions.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Receptor Tipo 1 de Angiotensina
/
Bloqueadores do Receptor Tipo 1 de Angiotensina II
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Avaliação Pré-Clínica de Medicamentos
Limite:
Humans
Idioma:
En
Revista:
Anal Bioanal Chem
Ano de publicação:
2018
Tipo de documento:
Article
País de afiliação:
China