{"database":"biostudies-literature","file_versions":[],"scores":null,"additional":{"omics_type":["Unknown"],"submitter":["Qiu X"],"funding":["Medical Research Council","Wellcome Trust"],"pubmed_abstract":["G protein-coupled receptors (GPCRs) belong to the most diverse group of membrane receptors with a conserved structure of seven transmembrane (TM) α-helices connected by intracellular and extracellular loops. Intracellular loop 3 (ICL3) connects TM5 and TM6, the two helices shown to play significant roles in receptor activation. Herein, we investigate the activation and signaling of the β<sub>1</sub> adrenergic receptor (β<sub>1</sub>AR) using mass spectrometry (MS) with a particular focus on the ICL3 loop. First, using native MS, we measure the extent of receptor coupling to an engineered Gα<sub>s</sub> subunit (mini G<sub>s</sub>) and show preferential coupling to β<sub>1</sub>AR with an intact ICL3 (β<sub>1</sub>AR_ICL3) compared to the truncated β<sub>1</sub>AR. Next, using hydrogen-deuterium exchange (HDX)-MS, we show how helix 5 of mini G<sub>s</sub> reports on the extent of receptor activation in the presence of a range of agonists. Then, exploring a range of solution conditions and using comparative HDX, we note additional HDX protection when ICL3 is present, implying that mini G<sub>s</sub> helix 5 presents a different binding conformation to the surface of β<sub>1</sub>AR_ICL3, a conclusion supported by MD simulation. Considering when this conformatonal change occurs we used time-resolved HDX and employed two functional assays to measure GDP release and cAMP production, with and without ICL3. We found that ICL3 exerts its effect on G<sub>s</sub> through enhanced cAMP production but does not affect GDP release. Together, our study uncovers potential roles of ICL3 in fine-tuning GPCR activation through subtle changes in the binding pose of helix 5, only after nucleotide release from G<sub>s</sub>."],"journal":["Journal of the American Chemical Society"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/S-EPMC11487556"],"repository":["biostudies-literature"],"pubmed_title":["Coupling and Activation of the β1 Adrenergic Receptor - The Role of the Third Intracellular Loop."],"pmcid":["PMC11487556"],"funding_grant_id":["221795/Z/20/Z","MR/T017961/1"],"pubmed_authors":["Chen YA","Rouse SL","Qiu X","Wang YQ","Chao K","Song S","Robinson CV","Yen HY"],"additional_accession":[]},"is_claimable":false,"name":"Coupling and Activation of the β1 Adrenergic Receptor - The Role of the Third Intracellular Loop.","description":"G protein-coupled receptors (GPCRs) belong to the most diverse group of membrane receptors with a conserved structure of seven transmembrane (TM) α-helices connected by intracellular and extracellular loops. Intracellular loop 3 (ICL3) connects TM5 and TM6, the two helices shown to play significant roles in receptor activation. Herein, we investigate the activation and signaling of the β<sub>1</sub> adrenergic receptor (β<sub>1</sub>AR) using mass spectrometry (MS) with a particular focus on the ICL3 loop. First, using native MS, we measure the extent of receptor coupling to an engineered Gα<sub>s</sub> subunit (mini G<sub>s</sub>) and show preferential coupling to β<sub>1</sub>AR with an intact ICL3 (β<sub>1</sub>AR_ICL3) compared to the truncated β<sub>1</sub>AR. Next, using hydrogen-deuterium exchange (HDX)-MS, we show how helix 5 of mini G<sub>s</sub> reports on the extent of receptor activation in the presence of a range of agonists. Then, exploring a range of solution conditions and using comparative HDX, we note additional HDX protection when ICL3 is present, implying that mini G<sub>s</sub> helix 5 presents a different binding conformation to the surface of β<sub>1</sub>AR_ICL3, a conclusion supported by MD simulation. Considering when this conformatonal change occurs we used time-resolved HDX and employed two functional assays to measure GDP release and cAMP production, with and without ICL3. We found that ICL3 exerts its effect on G<sub>s</sub> through enhanced cAMP production but does not affect GDP release. Together, our study uncovers potential roles of ICL3 in fine-tuning GPCR activation through subtle changes in the binding pose of helix 5, only after nucleotide release from G<sub>s</sub>.","dates":{"release":"2024-01-01T00:00:00Z","publication":"2024 Oct","modification":"2026-06-03T07:18:09.35Z","creation":"2025-04-06T09:34:51.222Z"},"accession":"S-EPMC11487556","cross_references":{"pubmed":["39359104"],"doi":["10.1021/jacs.4c11250"]}}