Control of conformational equilibria in the human B2 bradykinin receptor. Modeling of nonpeptidic ligand action and comparison to the rhodopsin structure.

Jean-Claude Bonnafous,Thierry Groblewski,Bernard Maigret,Jean-Luc Paquet,Philippe Vassault,Cecilia Poncé,Eric Richard,Christian Siatka,Jacky Marie,Renée Larguier,Didier Pruneau

doi:10.1074/jbc.m104875200

Abstract

A prototypic study of the molecular mechanisms of activation or inactivation of peptide hormone G protein-coupled receptors was carried out on the human B2 bradykinin receptor. A detailed pharmacological analysis of receptor mutants possessing either increased constitutive activity or impaired activation or ligand recognition allowed us to propose key residues participating in intramolecular interaction networks stabilizing receptor inactive or active conformations: Asn(113) and Tyr(115) (TM III), Trp(256) and Phe(259) (TM VI), Tyr(295) (TM VII) which are homologous of the rhodopsin residues Gly(120), Glu(122), Trp(265), Tyr(268), and Lys(296), respectively. An essential experimental finding was the spatial proximity between Asn(113), which is the cornerstone of inactive conformations, and Trp(256) which plays a subtle role in controlling the balance between active and inactive conformations. Molecular modeling and mutagenesis data showed that Trp(256) and Tyr(295) constitute, together with Gln(288), receptor contact points with original nonpeptidic ligands. It provided an explanation for the ligand inverse agonist behavior on the WT receptor, with underlying restricted motions of TMs III, VI, and VII, and its agonist behavior on the Ala(113) and Phe(256) constitutively activated mutants. These data on the B2 receptor emphasize that conformational equilibria are controlled in a coordinated fashion by key residues which are located at strategic positions for several G protein-coupled receptors. They are discussed in comparison with the recently determined rhodopsin crystallographic structure.

Highlights

A prototypic study of the molecular mechanisms of activation or inactivation of peptide hormone G proteincoupled receptors was carried out on the human B2 bradykinin receptor
A detailed pharmacological analysis of receptor mutants possessing either increased constitutive activity or impaired activation or ligand recognition allowed us to propose key residues participating in intramolecular interaction networks stabilizing receptor inactive or active conformations: Asn113 and Tyr115 (TM III), Trp256 and Phe259 (TM VI), Tyr295 (TM VII) which are homologous of the rhodopsin residues Gly120, Glu122, Trp265, Tyr268, and Lys296, respectively
These data on the B2 receptor emphasize that conformational equilibria are controlled in a coordinated fashion by key residues which are located at strategic positions for several G protein-coupled receptors

Summary

Introduction

A prototypic study of the molecular mechanisms of activation or inactivation of peptide hormone G proteincoupled receptors was carried out on the human B2 bradykinin receptor. Molecular modeling and mutagenesis data showed that Trp256 and Tyr295 constitute, together with Gln288, receptor contact points with original nonpeptidic ligands It provided an explanation for the ligand inverse agonist behavior on the WT receptor, with underlying restricted motions of TMs III, VI, and VII, and its agonist behavior on the Ala113 and Phe256 constitutively activated mutants. These data on the B2 receptor emphasize that conformational equilibria are controlled in a coordinated fashion by key residues which are located at strategic positions for several G protein-coupled receptors. Tivation phenomena have been widely exploited to dissect the intramolecular interaction networks which stabilize inactive receptor conformations [1], including random mutagenesis approaches to get deeper insight into the obvious complexity of these networks [2, 3]

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