Deciphering molecular determinants of GPBAR1-Gs protein interactions by HDX-MS and cryo-EM.

Fiche publication

Date publication

août 2025

Journal

Scientific reports

Auteurs

Membres identifiés du Cancéropôle Est :
Dr CIANFERANI Sarah

Tous les auteurs :
Castel J, Botzanowski T, Brooks I, Frechard A, Bey G, Schroeter M, Del Nero E, Debaene F, Ciesielski F, Zeyer D, Cianferani S, Morales R

Lien Pubmed

https://www.ncbi.nlm.nih.gov/pubmed/40858717

Résumé

Many physiological processes are dependent on G protein-coupled receptors (GPCRs), the biggest family of human membrane proteins and a significant class of therapeutic targets. Once activated by external stimuli, GPCRs use G proteins and arrestins as transducers to generate second messengers and trigger downstream signaling, leading to diverse signaling profiles. The G protein-coupled bile acid receptor 1 (GPBAR1, also known as Takeda G protein-coupled receptor 5, TGR5) is a class A bile acid membrane receptor that regulates energy homeostasis and glucose and lipid metabolism. GPBAR1/Gs protein interactions are implicated in the prevention of diabetes and the reduction of inflammatory responses, making GPBAR1 a potential therapeutic target for metabolic disorders. Here, we present combined hydrogen/deuterium exchange mass spectrometry (HDX-MS) and cryo-electron microscopy (cryo-EM) to identify the molecular determinants of GPBAR1 conformational dynamics upon G protein binding. Thanks to extensive optimization, we achieved over 75% sequence coverage by HDX-MS of a complete GPCR complex and a 2.5 Å resolution structure by cryo-EM, both of which are state-of-the-art. Altogether, our results provide information on the under-investigated GPBAR1 binding mode to its cognate G protein, pinpointing the synergic and powerful combination of higher cryo-EM and lower HDX-MS resolution techniques to dissect GPCR/G protein binding characteristics.