title: Structural Elements in the Gαs and Gαq C Termini That Mediate Selective G Protein-coupled Receptor (GPCR) Signaling.
abstract: Although the importance of the C terminus of the α subunit of the heterotrimeric G protein in G protein-coupled receptor (GPCR)-G protein pairing is well established, the structural basis of selective interactions remains unknown. Here, we combine live cell FRET-based measurements and molecular dynamics simulations of the interaction between the GPCR and a peptide derived from the C terminus of the Gα subunit (Gα peptide) to dissect the molecular mechanisms of G protein selectivity. We observe a direct link between Gα peptide binding and stabilization of the GPCR conformational ensemble. We find that cognate and non-cognate Gα peptides show deep and shallow binding, respectively, and in distinct orientations within the GPCR. Binding of the cognate Gα peptide stabilizes the agonist-bound GPCR conformational ensemble resulting in favorable binding energy and lower flexibility of the agonist-GPCR pair. We identify three hot spot residues (Gαs/Gαq-Gln-384/Leu-349, Gln-390/Glu-355, and Glu-392/Asn-357) that contribute to selective interactions between the β2-adrenergic receptor (β2-AR)-Gαs and V1A receptor (V1AR)-Gαq The Gαs and Gαq peptides adopt different orientations in β2-AR and V1AR, respectively. The β2-AR/Gαs peptide interface is dominated by electrostatic interactions, whereas the V1AR/Gαq peptide interactions are predominantly hydrophobic. Interestingly, our study reveals a role for both favorable and unfavorable interactions in G protein selection. Residue Glu-355 in Gαq prevents this peptide from interacting strongly with β2-AR. Mutagenesis to the Gαs counterpart (E355Q) imparts a cognate-like interaction. Overall, our study highlights the synergy in molecular dynamics and FRET-based approaches to dissect the structural basis of selective G protein interactions.
authors.affiliation: From the Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, Minnesota 55455.
authors.forename: Ansley
authors.initials: A
authors.lastname: Semack
authors.affiliation: the Department of Molecular Immunology, Beckman Research Institute of the City of Hope, Duarte, California 91010, and.
authors.forename: Manbir
authors.initials: M
authors.lastname: Sandhu
authors.affiliation: From the Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, Minnesota 55455, the Department of Cell and Developmental Biology, University of Michigan, Ann Arbor, Michigan 48109.
authors.forename: Rabia U
authors.initials: RU
authors.lastname: Malik
authors.affiliation: the Department of Molecular Immunology, Beckman Research Institute of the City of Hope, Duarte, California 91010, and.
authors.forename: Nagarajan
authors.initials: N
authors.lastname: Vaidehi
authors.affiliation: From the Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, Minnesota 55455, sivaraj@umn.edu.
authors.forename: Sivaraj
authors.initials: S
authors.lastname: Sivaramakrishnan
mesh_terms: D000818:Animals; D002460:Cell Line; D002864:Chromogranins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004795:Enzyme Stability; D031541:Fluorescence Resonance Energy Transfer; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051379:Mice; D056004:Molecular Dynamics Simulation; D020125:Mutation, Missense; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000072417:Protein Domains; D018343:Receptors, Adrenergic, beta-2; D034421:Sus scrofa
chemical_list: D002864:Chromogranins; C554061:GNAQ protein, human; D010455:Peptides; D018343:Receptors, Adrenergic, beta-2; C406732:GNAS protein, human; C498642:Gnas protein, mouse; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D019205:GTP-Binding Protein alpha Subunits, Gs
keywords: G protein; G protein-coupled receptor (GPCR); Receptor Conformation; cell signaling; fluorescence resonance energy transfer (FRET); molecular dynamics