uni-leipzig-open-access/json/s41467-023-37233-1

{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2023,12,10]],"date-time":"2023-12-10T12:49:42Z","timestamp":1702212582598},"reference-count":85,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2023,4,10]],"date-time":"2023-04-10T00:00:00Z","timestamp":1681084800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2023,4,10]],"date-time":"2023-04-10T00:00:00Z","timestamp":1681084800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"DOI":"10.13039\/100000009","name":"Foundation for the National Institutes of Health","doi-asserted-by":"publisher","award":["R01NS028471"]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Nat Commun"],"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Advances in structural biology have provided important mechanistic insights into signaling by the transmembrane core of G-protein coupled receptors (GPCRs); however, much less is known about intrinsically disordered regions such as the carboxyl terminus (CT), which is highly flexible and not visible in GPCR structures. The \u03b2<jats:sub>2<\/jats:sub>adrenergic receptor\u2019s (\u03b2<jats:sub>2<\/jats:sub>AR) 71 amino acid CT is a substrate for GPCR kinases and binds \u03b2-arrestins to regulate signaling. Here we show that the \u03b2<jats:sub>2<\/jats:sub>AR CT directly inhibits basal and agonist-stimulated signaling in cell lines lacking \u03b2-arrestins. Combining single-molecule fluorescence resonance energy transfer (FRET), NMR spectroscopy, and molecular dynamics simulations, we reveal that the negatively charged \u03b2<jats:sub>2<\/jats:sub>AR-CT serves as an autoinhibitory factor via interacting with the positively charged cytoplasmic surface of the receptor to limit access to G-proteins. The stability of this interaction is influenced by agonists and allosteric modulators, emphasizing that the CT plays important role in allosterically regulating GPCR activation.<\/jats:p>","DOI":"10.1038\/s41467-023-37233-1","type":"journal-article","created":{"date-parts":[[2023,4,10]],"date-time":"2023-04-10T09:03:20Z","timestamp":1681117400000},"update-policy":"http:\/\/dx.doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":7,"title":["Function and dynamics of the intrinsically disordered carboxyl terminus of \u03b22 adrenergic receptor"],"prefix":"10.1038","volume":"14","author":[{"given":"Jie","family":"Heng","sequence":"first","affiliation":[]},{"given":"Yunfei","family":"Hu","sequence":"additional","affiliation":[]},{"ORCID":"http:\/\/orcid.org\/0000-0002-9287-8704","authenticated-orcid":false,"given":"Guillermo","family":"P\u00e9rez-Hern\u00e1ndez","sequence":"additional","affiliation":[]},{"ORCID":"http:\/\/orcid.org\/0000-0003-0805-4049","authenticated-orcid":false,"given":"Asuka","family":"Inoue","sequence":"additional","affiliation":[]},{"given":"Jiawei","family":"Zhao","sequence":"additional","affiliation":[]},{"given":"Xiuyan","family":"Ma","sequence":"additional","affiliation":[]},{"given":"Xiaoou","family":"Sun","sequence":"additional","affiliation":[]},{"ORCID":"http:\/\/orcid.org\/0000-0002-2507-8619","authenticated-orcid":false,"given":"Kouki","family":"Kawakami","sequence":"additional","affiliation":[]},{"ORCID":"http:\/\/orcid.org\/0000-0002-8597-0609","authenticated-orcid":false,"given":"Tatsuya","family":"Ikuta","sequence":"additional","affiliation":[]},{"given":"Jienv","family":"Ding","sequence":"additional","affiliation":[]},{"given":"Yujie","family":"Yang","sequence":"additional","affiliation":[]},{"given":"Lujia","family":"Zhang","sequence":"additional","affiliation":[]},{"given":"Sijia","family":"Peng","sequence":"additional","affiliation":[]},{"given":"Xiaogang","family":"Niu","sequence":"additional","affiliation":[]},{"given":"Hongwei","family":"Li","sequence":"additional","affiliation":[]},{"ORCID":"http:\/\/orcid.org\/0000-0003-0397-9800","authenticated-orcid":false,"given":"Ramon","family":"Guix\u00e0-Gonz\u00e1lez","sequence":"additional","affiliation":[]},{"ORCID":"http:\/\/orcid.org\/0000-0002-1352-2540","authenticated-orcid":false,"given":"Changwen","family":"Jin","sequence":"additional","affiliation":[]},{"ORCID":"http:\/\/orcid.org\/0000-0003-0063-1104","authenticated-orcid":false,"given":"Peter W.","family":"Hildebrand","sequence":"additional","affiliation":[]},{"ORCID":"http:\/\/orcid.org\/0000-0002-0128-7766","authenticated-orcid":false,"given":"Chunlai","family":"Chen","sequence":"additional","affiliation":[]},{"ORCID":"http:\/\/orcid.org\/0000-0001-5958-3990","authenticated-orcid":false,"given":"Brian K.","family":"Kobilka","sequence":"additional","affiliation":[]}],"member":"297","published-online":{"date-parts":[[2023,4,10]]},"reference":[{"key":"37233_CR1","doi-asserted-by":"publisher","first-page":"45","DOI":"10.1038\/s41586-018-0259-z","volume":"559","author":"DM Thal","year":"2018","unstructured":"Thal, D. M., Glukhova, A., Sexton, P. M. & Christopoulos, A. Structural insights into G-protein-coupled receptor allostery. Nature 559, 45\u201353 (2018).","journal-title":"Nature"},{"key":"37233_CR2","doi-asserted-by":"publisher","first-page":"897","DOI":"10.1146\/annurev-biochem-060614-033910","volume":"87","author":"WI Weis","year":"2018","unstructured":"Weis, W. I. & Kobilka, B. K. The Molecular Basis of G-protein-Coupled Receptor Activation. Annu Rev. Biochem. 87, 897\u2013919 (2018).","journal-title":"Annu Rev. Biochem."},{"key":"37233_CR3","doi-asserted-by":"publisher","first-page":"198","DOI":"10.1126\/science.1169377","volume":"324","author":"RG Smock","year":"2009","unstructured":"Smock, R. G. & Gierasch, L. M. Sending Signals Dynamically. Science 324, 198\u2013203 (2009).","journal-title":"Science"},{"key":"37233_CR4","doi-asserted-by":"publisher","first-page":"18","DOI":"10.1038\/nrm3920","volume":"16","author":"PE Wright","year":"2015","unstructured":"Wright, P. E. & Dyson, H. J. Intrinsically disordered proteins in cellular signaling and regulation. Nat. Rev. Mol. Cell Biol. 16, 18\u201329 (2015).","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"37233_CR5","doi-asserted-by":"publisher","first-page":"103","DOI":"10.1093\/protein\/gzi004","volume":"18","author":"V-P Jaakola","year":"2005","unstructured":"Jaakola, V.-P., Prilusky, J., Sussman, J. L. & Goldman, A. G-protein-coupled receptors show unusual patterns of intrinsic unfolding. Protein Eng. Des. Sel. 18, 103\u2013110 (2005).","journal-title":"Protein Eng. Des. Sel."},{"key":"37233_CR6","doi-asserted-by":"publisher","first-page":"129","DOI":"10.1016\/j.sbi.2014.08.002","volume":"27","author":"AJ Venkatakrishnan","year":"2014","unstructured":"Venkatakrishnan, A. J. et al. Structured and disordered facets of the GPCR fold. Curr. Opin. Struct. Biol. 27, 129\u2013137 (2014).","journal-title":"Curr. Opin. Struct. Biol."},{"key":"37233_CR7","doi-asserted-by":"publisher","first-page":"1717","DOI":"10.1111\/j.1476-5381.2011.01552.x","volume":"165","author":"AC Magalhaes","year":"2012","unstructured":"Magalhaes, A. C., Dunn, H. & Ferguson, S. S. Regulation of GPCR activity, trafficking and localization by GPCR-interacting proteins. Br. J. Pharmacol. 165, 1717\u20131736 (2012).","journal-title":"Br. J. Pharmacol."},{"key":"37233_CR8","doi-asserted-by":"publisher","unstructured":"Marti-Solano, M. et al. Combinatorial expression of GPCR isoforms affects signaling and drug responses. Nature 1\u201328 https:\/\/doi.org\/10.1038\/s41586-020-2888-2 (2020).","DOI":"10.1038\/s41586-020-2888-2"},{"key":"37233_CR9","doi-asserted-by":"publisher","first-page":"8944","DOI":"10.1074\/jbc.270.15.8944","volume":"270","author":"J Alblas","year":"1995","unstructured":"Alblas, J., Etten, I., van, Khanum, A. & Moolenaar, W. H. C-terminal truncation of the neurokinin-2 receptor causes enhanced and sustained agonist-induced signaling. Role of receptor phosphorylation in signal attenuation. J. Biol. Chem. 270, 8944\u20138951 (1995).","journal-title":"J. Biol. Chem."},{"key":"37233_CR10","doi-asserted-by":"publisher","first-page":"9987","DOI":"10.1016\/S0021-9258(18)92917-0","volume":"266","author":"EM Parker","year":"1991","unstructured":"Parker, E. M. & Ross, E. M. Truncation of the extended carboxyl-terminal domain increases the expression and regulatory activity of the avian beta-adrenergic receptor. J. Biol. Chem. 266, 9987\u20139996 (1991).","journal-title":"J. Biol. Chem."},{"key":"37233_CR11","doi-asserted-by":"publisher","first-page":"1857","DOI":"10.1074\/jbc.271.4.1857","volume":"271","author":"H Hasegawa","year":"1996","unstructured":"Hasegawa, H., Negishi, M. & Ichikawa, A. Two isoforms of the prostaglandin E receptor EP3 subtype different in agonist-independent constitutive activity. J. Biol. Chem. 271, 1857\u20131860 (1996).","journal-title":"J. Biol. Chem."},{"key":"37233_CR12","doi-asserted-by":"publisher","first-page":"1041","DOI":"10.1074\/jbc.270.3.1041","volume":"270","author":"N Matus-Leibovitch","year":"1995","unstructured":"Matus-Leibovitch, N., Nussenzveig, D. R., Gershengorn, M. C. & Oron, Y. Truncation of the thyrotropin-releasing hormone receptor carboxyl tail causes constitutive activity and leads to impaired responsiveness in Xenopus oocytes and AtT20 cells. J. Biol. Chem. 270, 1041\u20131047 (1995).","journal-title":"J. Biol. Chem."},{"key":"37233_CR13","doi-asserted-by":"publisher","first-page":"30283","DOI":"10.1074\/jbc.M212918200","volume":"278","author":"H Pankevych","year":"2003","unstructured":"Pankevych, H., Korkhov, V., Freissmuth, M. & Nanoff, C. Truncation of the A1 adenosine receptor reveals distinct roles of the membrane-proximal carboxyl terminus in receptor folding and G-protein coupling. J. Biol. Chem. 278, 30283\u201330293 (2003).","journal-title":"J. Biol. Chem."},{"key":"37233_CR14","doi-asserted-by":"publisher","first-page":"1101","DOI":"10.1016\/j.cell.2015.04.043","volume":"161","author":"A Manglik","year":"2015","unstructured":"Manglik, A. et al. Structural Insights into the Dynamic Process of \u03b22-Adrenergic Receptor Signaling. Cell 161, 1101\u20131111 (2015).","journal-title":"Cell"},{"key":"37233_CR15","doi-asserted-by":"publisher","first-page":"68","DOI":"10.1038\/nature22354","volume":"547","author":"GG Gregorio","year":"2017","unstructured":"Gregorio, G. G. et al. Single-molecule analysis of ligand efficacy in \u03b22AR-G-protein activation. Nature 547, 68\u201373 (2017).","journal-title":"Nature"},{"key":"37233_CR16","doi-asserted-by":"publisher","first-page":"171","DOI":"10.1038\/nmeth742","volume":"2","author":"C Hoffmann","year":"2005","unstructured":"Hoffmann, C. et al. A FlAsH-based FRET approach to determine G-protein-coupled receptor activation in living cells. Nat Methods. 2, 171\u2013176 (2005).","journal-title":"Nat Methods."},{"key":"37233_CR17","doi-asserted-by":"publisher","first-page":"36188","DOI":"10.1074\/jbc.M110.175604","volume":"285","author":"S Reiner","year":"2010","unstructured":"Reiner, S., Ambrosio, M. & Hoffmann, C. & lohse, martin j. Differential Signaling of the Endogenous Agonists at the beta(2)-Adrenergic Receptor. J. Biol. Chem. 285, 36188\u201336198 (2010).","journal-title":"J. Biol. Chem."},{"key":"37233_CR18","doi-asserted-by":"publisher","first-page":"159","DOI":"10.1016\/j.tips.2007.12.002","volume":"29","author":"MJ lohse","year":"2008","unstructured":"lohse, M. J. et al. Optical techniques to analyze real-time activation and signaling of G-protein-coupled receptors. Trends Pharmacol. Sci. 29, 159\u2013165 (2008).","journal-title":"Trends Pharmacol. Sci."},{"key":"37233_CR19","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-017-02632-8","volume":"9","author":"Y Shiraishi","year":"2018","unstructured":"Shiraishi, Y. et al. Phosphorylation-induced conformation of \u03b22-adrenoceptor related to arrestin recruitment revealed by NMR. Nat. Commun. 9, 194 (2018).","journal-title":"Nat. Commun."},{"key":"37233_CR20","first-page":"577","volume":"60","author":"E Devic","year":"2001","unstructured":"Devic, E., Xiang, Y., Gould, D. & Kobilka, B. Beta-adrenergic receptor subtype-specific signaling in cardiac myocytes from beta(1) and beta(2) adrenoceptor knockout mice. Mol. Pharmacol. 60, 577\u2013583 (2001).","journal-title":"Mol. Pharmacol."},{"key":"37233_CR21","doi-asserted-by":"publisher","first-page":"7094","DOI":"10.1016\/S0021-9258(18)88892-5","volume":"260","author":"JL Benovic","year":"1985","unstructured":"Benovic, J. L. et al. Phosphorylation of the mammalian beta-adrenergic receptor by cyclic AMP-dependent protein kinase. Regulation of the rate of receptor phosphorylation and dephosphorylation by agonist occupancy and effects on coupling of the receptor to the stimulatory guanine nucleotide regulatory protein. J. Biol. Chem. 260, 7094\u20137101 (1985).","journal-title":"J. Biol. Chem."},{"key":"37233_CR22","doi-asserted-by":"publisher","first-page":"3106","DOI":"10.1016\/S0021-9258(18)61475-9","volume":"262","author":"M Bouvier","year":"1987","unstructured":"Bouvier, M., Leeb-Lundberg, L. M., Benovic, J. L., Caron, M. G. & Lefkowitz, R. J. Regulation of adrenergic receptor function by phosphorylation. II. Effects of agonist occupancy on phosphorylation of alpha 1- and beta 2-adrenergic receptors by protein kinase C and the cyclic AMP-dependent protein kinase. J. Biol. Chem. 262, 3106\u20133113 (1987).","journal-title":"J. Biol. Chem."},{"key":"37233_CR23","doi-asserted-by":"publisher","first-page":"2797","DOI":"10.1073\/pnas.83.9.2797","volume":"83","author":"JL Benovic","year":"1986","unstructured":"Benovic, J. L., Strasser, R. H., Caron, M. G. & Lefkowitz, R. J. Beta-adrenergic receptor kinase: identification of a novel protein kinase that phosphorylates the agonist-occupied form of the receptor. Proc. Natl Acad. Sci. 83, 2797\u20132801 (1986).","journal-title":"Proc. Natl Acad. Sci."},{"key":"37233_CR24","doi-asserted-by":"publisher","first-page":"ra51","DOI":"10.1126\/scisignal.2001707","volume":"4","author":"KN Nobles","year":"2011","unstructured":"Nobles, K. N. et al. Distinct phosphorylation sites on the \u03b2(2)-adrenergic receptor establish a barcode that encodes differential functions of \u03b2-arrestin. Sci. Signal. 4, ra51\u2013ra51 (2011).","journal-title":"Sci. Signal."},{"key":"37233_CR25","doi-asserted-by":"publisher","first-page":"1547","DOI":"10.1126\/science.2163110","volume":"248","author":"MJ Lohse","year":"1990","unstructured":"Lohse, M. J., Benovic, J. L., Codina, J., Caron, M. G. & Lefkowitz, R. J. beta-Arrestin: a protein that regulates beta-adrenergic receptor function. Science 248, 1547\u20131550 (1990).","journal-title":"Science"},{"key":"37233_CR26","doi-asserted-by":"publisher","first-page":"549","DOI":"10.1038\/nature10361","volume":"477","author":"SGF Rasmussen","year":"2011","unstructured":"Rasmussen, S. G. F. et al. Crystal structure of the \u03b22 adrenergic receptor-Gs protein complex. Nature 477, 549\u2013555 (2011).","journal-title":"Nature"},{"key":"37233_CR27","doi-asserted-by":"publisher","first-page":"23096","DOI":"10.1073\/pnas.2009786117","volume":"117","author":"X Ma","year":"2020","unstructured":"Ma, X. et al. Analysis of \u03b22AR-Gs and \u03b22AR-Gi complex formation by NMR spectroscopy. Proc. Natl Acad. Sci. 117, 23096\u201323105 (2020).","journal-title":"Proc. Natl Acad. Sci."},{"key":"37233_CR28","doi-asserted-by":"publisher","first-page":"317","DOI":"10.1038\/nature22070","volume":"545","author":"T Flock","year":"2017","unstructured":"Flock, T. et al. Selectivity determinants of GPCR-G-protein binding. Nature 545, 317\u2013322 (2017).","journal-title":"Nature"},{"key":"37233_CR29","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1159\/000068914","volume":"12","author":"SK-F Wong","year":"2003","unstructured":"Wong, S. K.-F. G protein selectivity is regulated by multiple intracellular regions of GPCRs. Neurosignals 12, 1\u201312 (2003).","journal-title":"Neurosignals"},{"key":"37233_CR30","doi-asserted-by":"publisher","first-page":"407","DOI":"10.1016\/j.cell.2017.03.047","volume":"169","author":"KE Komolov","year":"2017","unstructured":"Komolov, K. E. et al. Structural and Functional Analysis of a \u03b22-Adrenergic Receptor Complex with GRK5. Cell 169, 407\u2013421.e16 (2017).","journal-title":"Cell"},{"key":"37233_CR31","doi-asserted-by":"publisher","first-page":"31840","DOI":"10.1074\/jbc.M806086200","volume":"283","author":"C Krasel","year":"2008","unstructured":"Krasel, C. et al. Dual Role of the 2-Adrenergic Receptor C Terminus for the Binding of -Arrestin and Receptor Internalization. J. Biol. Chem. 283, 31840\u201331848 (2008).","journal-title":"J. Biol. Chem."},{"key":"37233_CR32","doi-asserted-by":"publisher","first-page":"447","DOI":"10.1038\/383447a0","volume":"383","author":"OB Goodman Jr","year":"1996","unstructured":"Goodman, O. B. Jr et al. \u03b2-Arrestin acts as a clathrin adaptor in endocytosis of the \u03b22-adrenergic receptor. Nature 383, 447\u2013450 (1996).","journal-title":"Nature"},{"key":"37233_CR33","doi-asserted-by":"publisher","first-page":"32248","DOI":"10.1074\/jbc.274.45.32248","volume":"274","author":"RH Oakley","year":"1999","unstructured":"Oakley, R. H., Laporte, S. A., Holt, J. A., Barak, L. S. & Caron, M. G. Association of beta-arrestin with G-protein-coupled receptors during clathrin-mediated endocytosis dictates the profile of receptor resensitization. J. Biol. Chem. 274, 32248\u201332257 (1999).","journal-title":"J. Biol. Chem."},{"key":"37233_CR34","doi-asserted-by":"publisher","first-page":"819","DOI":"10.1038\/nrm2803","volume":"10","author":"SL Ritter","year":"2009","unstructured":"Ritter, S. L. & Hall, R. A. Fine-tuning of GPCR activity by receptor-interacting proteins. Nat. Rev. Mol. Cell Biol. 10, 819\u2013830 (2009).","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"37233_CR35","doi-asserted-by":"publisher","first-page":"286","DOI":"10.1038\/45816","volume":"401","author":"TT Cao","year":"1999","unstructured":"Cao, T. T., Deacon, H. W., Reczek, D., Bretscher, A. & Zastrow, Mvon A kinase-regulated PDZ-domain interaction controls endocytic sorting of the beta2-adrenergic receptor. Nature 401, 286\u2013290 (1999).","journal-title":"Nature"},{"key":"37233_CR36","doi-asserted-by":"publisher","first-page":"45145","DOI":"10.1074\/jbc.M106087200","volume":"276","author":"M Cong","year":"2001","unstructured":"Cong, M. et al. Binding of the beta2 adrenergic receptor to N-ethylmaleimide-sensitive factor regulates receptor recycling. J. Biol. Chem. 276, 45145\u201345152 (2001).","journal-title":"J. Biol. Chem."},{"key":"37233_CR37","doi-asserted-by":"publisher","first-page":"626","DOI":"10.1038\/33458","volume":"392","author":"RA Hall","year":"1998","unstructured":"Hall, R. A. et al. The beta2-adrenergic receptor interacts with the Na+\/H+-exchanger regulatory factor to control Na+\/H+ exchange. Nature 392, 626\u2013630 (1998).","journal-title":"Nature"},{"key":"37233_CR38","doi-asserted-by":"publisher","first-page":"12785","DOI":"10.1074\/jbc.M110.203091","volume":"286","author":"K Xiao","year":"2011","unstructured":"Xiao, K. & Shenoy, S. K. Beta2-adrenergic receptor lysosomal trafficking is regulated by ubiquitination of lysyl residues in two distinct receptor domains. J. Biol. Chem. 286, 12785\u201312795 (2011).","journal-title":"J. Biol. Chem."},{"key":"37233_CR39","doi-asserted-by":"publisher","first-page":"1077","DOI":"10.1002\/pro.2494","volume":"23","author":"V Pejaver","year":"2014","unstructured":"Pejaver, V. et al. The structural and functional signatures of proteins that undergo multiple events of post-translational modification. Protein Sci. 23, 1077\u20131093 (2014).","journal-title":"Protein Sci."},{"key":"37233_CR40","doi-asserted-by":"publisher","first-page":"120","DOI":"10.1124\/pharmrev.120.000082","volume":"73","author":"A Patwardhan","year":"2021","unstructured":"Patwardhan, A., Cheng, N. & Trejo, J. Post-Translational Modifications of G-protein-Coupled Receptors Control Cellular Signaling Dynamics in Space and Time. Pharmacol. Rev. 73, 120\u2013151 (2021).","journal-title":"Pharmacol. Rev."},{"key":"37233_CR41","doi-asserted-by":"publisher","first-page":"17566","DOI":"10.1021\/jacs.1c07369","volume":"143","author":"Y Li","year":"2021","unstructured":"Li, Y. et al. Chemical Synthesis of a Full-Length G-Protein-Coupled Receptor \u03b22-Adrenergic Receptor with Defined Modification Patterns at the C-Terminus. J. Am. Chem. Soc. 143, 17566\u201317576 (2021).","journal-title":"J. Am. Chem. Soc."},{"key":"37233_CR42","doi-asserted-by":"publisher","first-page":"2562","DOI":"10.1073\/pnas.1701529114","volume":"114","author":"TJI Cahill","year":"2017","unstructured":"Cahill, T. J. I. et al. Distinct conformations of GPCR-beta-arrestin complexes mediate desensitization, signaling, and endocytosis. Proc. Natl Acad. Sci. 114, 2562\u20132567 (2017).","journal-title":"Proc. Natl Acad. Sci."},{"key":"37233_CR43","doi-asserted-by":"publisher","first-page":"603","DOI":"10.3390\/biomedicines8120603","volume":"8","author":"AR Jain","year":"2020","unstructured":"Jain, A. R., McGraw, C. & Robinson, A. S. The Specificity of Downstream Signaling for A1 and A2AR Does Not Depend on the C-Terminus. Biomedicines 8, 603 (2020).","journal-title":"Biomedicines"},{"key":"37233_CR44","doi-asserted-by":"publisher","first-page":"1933","DOI":"10.1016\/j.cell.2019.04.044","volume":"177","author":"A Inoue","year":"2019","unstructured":"Inoue, A. et al. Illuminating G-Protein-Coupling Selectivity of GPCRs. Cell 177, 1933\u20131947.e25 (2019).","journal-title":"Cell"},{"key":"37233_CR45","doi-asserted-by":"publisher","first-page":"13895","DOI":"10.1074\/jbc.M611904200","volume":"282","author":"S Granier","year":"2007","unstructured":"Granier, S. et al. Structure and conformational changes in the C-terminal domain of the beta2-adrenoceptor: insights from fluorescence resonance energy transfer studies. J. Biol. Chem. 282, 13895\u201313905 (2007).","journal-title":"J. Biol. Chem."},{"key":"37233_CR46","doi-asserted-by":"publisher","first-page":"575","DOI":"10.1038\/nature12572","volume":"502","author":"AM Ring","year":"2013","unstructured":"Ring, A. M. et al. Adrenaline-activated structure of \u03b22-adrenoceptor stabilized by an engineered nanobody. Nature 502, 575\u2013579 (2013).","journal-title":"Nature"},{"key":"37233_CR47","doi-asserted-by":"publisher","unstructured":"Liu, X. et al. Mechanism of intracellular allosteric \u03b22AR antagonist revealed by X-ray crystal structure. Nature. 1\u201319 https:\/\/doi.org\/10.1038\/nature23652 (2017).","DOI":"10.1038\/nature23652"},{"key":"37233_CR48","doi-asserted-by":"publisher","first-page":"1283","DOI":"10.1126\/science.aaw8981","volume":"364","author":"X Liu","year":"2019","unstructured":"Liu, X. et al. Mechanism of \u03b22AR regulation by an intracellular positive allosteric modulator. Science 364, 1283\u20131287 (2019).","journal-title":"Science"},{"key":"37233_CR49","doi-asserted-by":"publisher","first-page":"749","DOI":"10.1038\/s41589-020-0549-2","volume":"16","author":"X Liu","year":"2020","unstructured":"Liu, X. et al. An allosteric modulator binds to a conformational hub in the \u03b22 adrenergic receptor. Nat. Chem. Biol. 16, 749\u2013755 (2020).","journal-title":"Nat. Chem. Biol."},{"key":"37233_CR50","doi-asserted-by":"publisher","first-page":"e0175642","DOI":"10.1371\/journal.pone.0175642","volume":"12","author":"R Nehm\u00e9","year":"2017","unstructured":"Nehm\u00e9, R. et al. Mini-G-proteins: Novel tools for studying GPCRs in their active conformation. PLoS ONE. 12, e0175642 (2017).","journal-title":"PLoS ONE."},{"key":"37233_CR51","doi-asserted-by":"publisher","first-page":"1232","DOI":"10.1016\/j.cell.2019.04.022","volume":"177","author":"Y Du","year":"2019","unstructured":"Du, Y. et al. Assembly of a GPCR-G-protein Complex. Cell 177, 1232\u20131242.e11 (2019).","journal-title":"Cell"},{"key":"37233_CR52","doi-asserted-by":"publisher","first-page":"1243","DOI":"10.1016\/j.cell.2019.04.021","volume":"177","author":"X Liu","year":"2019","unstructured":"Liu, X. et al. Structural Insights into the Process of GPCR-G-protein Complex Formation. Cell 177, 1243\u20131251.e12 (2019).","journal-title":"Cell"},{"key":"37233_CR53","doi-asserted-by":"publisher","first-page":"1941","DOI":"10.1529\/biophysj.106.082487","volume":"91","author":"SA McKinney","year":"2006","unstructured":"McKinney, S. A., Joo, C. & Ha, T. Analysis of single-molecule FRET trajectories using hidden Markov modeling. Biophysj 91, 1941\u20131951 (2006).","journal-title":"Biophysj"},{"key":"37233_CR54","doi-asserted-by":"publisher","first-page":"1048","DOI":"10.1111\/j.1476-5381.2010.00754.x","volume":"160","author":"JG Baker","year":"2010","unstructured":"Baker, J. G. The selectivity of \u03b2-adrenoceptor agonists at human \u03b21-, \u03b22- and \u03b23-adrenoceptors. Br. J. Pharmacol. 160, 1048\u20131061 (2010).","journal-title":"Br. J. Pharmacol."},{"key":"37233_CR55","doi-asserted-by":"publisher","first-page":"571","DOI":"10.1016\/j.jmb.2004.07.044","volume":"342","author":"T Okada","year":"2004","unstructured":"Okada, T. et al. The retinal conformation and its environment in rhodopsin in light of a new 2.2 angstrom crystal structure. J. Mol. Biol. 342, 571\u2013583 (2004).","journal-title":"J. Mol. Biol."},{"key":"37233_CR56","first-page":"363","volume":"453","author":"M Murakami","year":"2008","unstructured":"Murakami, M. & Kouyama, T. Cryst. Struct. Squid Rhodopsin. 453, 363\u2013367 (2008).","journal-title":"Cryst. Struct. Squid Rhodopsin."},{"key":"37233_CR57","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1021\/bi011870b","volume":"41","author":"ERP Zuiderweg","year":"2002","unstructured":"Zuiderweg, E. R. P. Mapping protein\u2212protein interactions in solution by NMR spectroscopy \u2020. Biochemistry 41, 1\u20137 (2002).","journal-title":"Biochemistry"},{"key":"37233_CR58","doi-asserted-by":"publisher","first-page":"3","DOI":"10.1007\/s10858-010-9464-2","volume":"49","author":"MA Refaei","year":"2011","unstructured":"Refaei, M. A. et al. Observing selected domains in multi-domain proteins via sortase-mediated ligation and NMR spectroscopy. J. Biomolecular NMR. 49, 3\u20137 (2011).","journal-title":"J. Biomolecular NMR."},{"key":"37233_CR59","doi-asserted-by":"crossref","unstructured":"Rasmussen, S. G. F. et al. Structure of a nanobody-stabilized active state of the \u03b22 adrenoceptor. Nature 469, 175\u2013180 (2010).","DOI":"10.1038\/nature09648"},{"key":"37233_CR60","doi-asserted-by":"publisher","first-page":"4108","DOI":"10.1021\/cr900033p","volume":"109","author":"GM Clore","year":"2009","unstructured":"Clore, G. M. & Iwahara, J. Theory, Practice, and Applications of Paramagnetic Relaxation Enhancement for the Characterization of Transient Low-Population States of Biological Macromolecules and Their Complexes. Chem. Rev. 109, 4108\u20134139 (2009).","journal-title":"Chem. Rev."},{"key":"37233_CR61","doi-asserted-by":"publisher","first-page":"332","DOI":"10.1016\/j.str.2012.12.013","volume":"21","author":"T Trudeau","year":"2013","unstructured":"Trudeau, T. et al. Structure and intrinsic disorder in protein autoinhibition. Structure 21, 332\u2013341 (2013).","journal-title":"Structure"},{"key":"37233_CR62","doi-asserted-by":"publisher","first-page":"409","DOI":"10.1016\/S0960-9822(00)00419-X","volume":"10","author":"IDC Fraser","year":"2000","unstructured":"Fraser, I. D. C. et al. Assembly of an A kinase-anchoring protein\u2013\u03b22-adrenergic receptor complex facilitates receptor phosphorylation and signaling. Curr. Biol. 10, 409\u2013412 (2000).","journal-title":"Curr. Biol."},{"key":"37233_CR63","doi-asserted-by":"publisher","first-page":"1123","DOI":"10.1038\/s41594-019-0330-y","volume":"26","author":"AH Nguyen","year":"2019","unstructured":"Nguyen, A. H. et al. Structure of an endosomal signaling GPCR-G-protein-\u03b2-arrestin megacomplex. Nat. Struct. Mol. Biol. 26, 1123\u20131131 (2019).","journal-title":"Nat. Struct. Mol. Biol."},{"key":"37233_CR64","doi-asserted-by":"publisher","first-page":"400","DOI":"10.1021\/acschembio.5b00753","volume":"11","author":"AS Dixon","year":"2016","unstructured":"Dixon, A. S. et al. NanoLuc Complementation Reporter Optimized for Accurate Measurement of Protein Interactions in Cells. Acs Chem. Biol. 11, 400\u2013408 (2016).","journal-title":"Acs Chem. Biol."},{"key":"37233_CR65","doi-asserted-by":"publisher","first-page":"e46039","DOI":"10.1371\/journal.pone.0046039","volume":"7","author":"Y Zou","year":"2012","unstructured":"Zou, Y., Weis, W. I. & Kobilka, B. K. N-Terminal T4 Lysozyme Fusion Facilitates Crystallization of a G-protein Coupled Receptor. Plos One. 7, e46039 (2012).","journal-title":"Plos One."},{"key":"37233_CR66","first-page":"201722336","volume":"56","author":"DP Staus","year":"2018","unstructured":"Staus, D. P. et al. Sortase ligation enables homogeneous GPCR phosphorylation to reveal diversity in \u03b2-arrestin coupling. Proc. Natl Acad. Sci. 56, 201722336 (2018).","journal-title":"Proc. Natl Acad. Sci."},{"key":"37233_CR67","doi-asserted-by":"publisher","first-page":"372","DOI":"10.1016\/j.celrep.2017.12.048","volume":"22","author":"M Yang","year":"2018","unstructured":"Yang, M. et al. The Conformational Dynamics of Cas9 Governing DNA Cleavage Are Revealed by Single-Molecule FRET. Cell Rep. 22, 372\u2013382 (2018).","journal-title":"Cell Rep."},{"key":"37233_CR68","doi-asserted-by":"publisher","first-page":"gky636","DOI":"10.1093\/nar\/gky636","volume":"46","author":"B Wu","year":"2018","unstructured":"Wu, B. et al. Translocation kinetics and structural dynamics of ribosomes are modulated by the conformational plasticity of downstream pseudoknots. Nucl. Acids Res. 46, gky636 (2018).","journal-title":"Nucl. Acids Res."},{"key":"37233_CR69","doi-asserted-by":"publisher","first-page":"367","DOI":"10.1016\/j.molcel.2011.03.024","volume":"42","author":"C Chen","year":"2011","unstructured":"Chen, C. et al. Single-Molecule Fluorescence Measurements of Ribosomal Translocation Dynamics. Mol. Cell. 42, 367\u2013377 (2011).","journal-title":"Mol. Cell."},{"key":"37233_CR70","doi-asserted-by":"publisher","first-page":"277","DOI":"10.1007\/BF00197809","volume":"6","author":"F Delaglio","year":"1995","unstructured":"Delaglio, F. et al. NMRPipe: A multidimensional spectral processing system based on UNIX pipes. J. Biomol. Nmr. 6, 277\u2013293 (1995).","journal-title":"J. Biomol. Nmr."},{"key":"37233_CR71","first-page":"313","volume":"278","author":"AK Downing","year":"2004","unstructured":"Downing, A. K. & Johnson, B. A. Protein NMR Techniques. Methods Mol. Biol. Clifton N. J. 278, 313\u2013352 (2004).","journal-title":"Methods Mol. Biol. Clifton N. J."},{"key":"37233_CR72","doi-asserted-by":"publisher","first-page":"9501","DOI":"10.1073\/pnas.0811437106","volume":"106","author":"XJ Yao","year":"2009","unstructured":"Yao, X. J. et al. The effect of ligand efficacy on the formation and stability of a GPCR-G-protein complex. Proc. Natl Acad. Sci. 106, 9501\u20139506 (2009).","journal-title":"Proc. Natl Acad. Sci."},{"key":"37233_CR73","doi-asserted-by":"publisher","first-page":"33","DOI":"10.1016\/0263-7855(96)00018-5","volume":"14","author":"W Humphrey","year":"1996","unstructured":"Humphrey, W., Dalke, A. & Schulten, K. VMD: Visual molecular dynamics. J. Mol. Graph. 14, 33\u201338 (1996).","journal-title":"J. Mol. Graph."},{"key":"37233_CR74","doi-asserted-by":"publisher","first-page":"gky427","DOI":"10.1093\/nar\/gky427","volume":"46","author":"A Waterhouse","year":"2018","unstructured":"Waterhouse, A. et al. SWISS-MODEL: homology modelling of protein structures and complexes. Nucl. Acids Res. 46, gky427 (2018).","journal-title":"Nucl. Acids Res."},{"key":"37233_CR75","doi-asserted-by":"publisher","first-page":"229","DOI":"10.1002\/prot.24179","volume":"81","author":"D Xu","year":"2013","unstructured":"Xu, D. & Zhang, Y. Toward optimal fragment generations for ab initio protein structure assembly. Proteins Struct. Funct. Bioinform. 81, 229\u2013239 (2013).","journal-title":"Proteins Struct. Funct. Bioinform."},{"key":"37233_CR76","doi-asserted-by":"publisher","first-page":"gkaa440","DOI":"10.1093\/nar\/gkaa440","volume":"48","author":"E Mayol","year":"2020","unstructured":"Mayol, E. et al. HomolWat: a web server tool to incorporate \u2018homologous\u2019 water molecules into GPCR structures. Nucl. Acids Res. 48, gkaa440 (2020).","journal-title":"Nucl. Acids Res."},{"key":"37233_CR77","doi-asserted-by":"publisher","first-page":"e880","DOI":"10.1371\/journal.pone.0000880","volume":"2","author":"S Jo","year":"2007","unstructured":"Jo, S., Kim, T. & Im, W. Automated Builder and Database of Protein\/Membrane Complexes for Molecular Dynamics Simulations. Plos One. 2, e880 (2007).","journal-title":"Plos One."},{"key":"37233_CR78","doi-asserted-by":"publisher","first-page":"19","DOI":"10.1016\/j.softx.2015.06.001","volume":"1","author":"MJ Abraham","year":"2015","unstructured":"Abraham, M. J. et al. GROMACS: High performance molecular simulations through multi-level parallelism from laptops to supercomputers. Softwarex 1, 19\u201325 (2015).","journal-title":"Softwarex"},{"key":"37233_CR79","doi-asserted-by":"publisher","first-page":"71","DOI":"10.1038\/nmeth.4067","volume":"14","author":"J Huang","year":"2017","unstructured":"Huang, J. et al. CHARMM36m: an improved force field for folded and intrinsically disordered proteins. Nat. Methods. 14, 71\u201373 (2017).","journal-title":"Nat. Methods."},{"key":"37233_CR80","doi-asserted-by":"crossref","first-page":"671","DOI":"10.1002\/jcc.21367","volume":"31","author":"K Vanommeslaeghe","year":"2010","unstructured":"Vanommeslaeghe, K. et al. CHARMM general force field: A force field for drug\u2010like molecules compatible with the CHARMM all\u2010atom additive biological force fields. J. Comput Chem. 31, 671\u2013690 (2010).","journal-title":"J. Comput Chem."},{"key":"37233_CR81","doi-asserted-by":"publisher","first-page":"244119","DOI":"10.1063\/1.5053582","volume":"149","author":"E Hruska","year":"2018","unstructured":"Hruska, E., Abella, J. R., N\u00fcske, F., Kavraki, L. E. & Clementi, C. Quantitative comparison of adaptive sampling methods for protein dynamics. J. Chem. Phys. 149, 244119 (2018).","journal-title":"J. Chem. Phys."},{"key":"37233_CR82","doi-asserted-by":"publisher","first-page":"5747","DOI":"10.1021\/acs.jctc.5b00737","volume":"11","author":"MI Zimmerman","year":"2015","unstructured":"Zimmerman, M. I. & Bowman, G. R. FAST Conformational Searches by Balancing Exploration\/Exploitation Trade-Offs. J. Chem. Theory Comput. 11, 5747\u20135757 (2015).","journal-title":"J. Chem. Theory Comput."},{"key":"37233_CR83","doi-asserted-by":"publisher","first-page":"1528","DOI":"10.1016\/j.bpj.2015.08.015","volume":"109","author":"RT McGibbon","year":"2015","unstructured":"McGibbon, R. T. et al. MDTraj: A Modern Open Library for the Analysis of Molecular Dynamics Trajectories. Biophys. J. 109, 1528\u20131532 (2015).","journal-title":"Biophys. J."},{"key":"37233_CR84","doi-asserted-by":"crossref","unstructured":"P\u00e9rez-Hern\u00e1ndez G. & Hildebrand P-. W. mdciao: Accessible Analysis and Visualization of Molecular Dynamics Simulation Data. Preprint at https:\/\/www.biorxiv.org\/content\/10.1101\/2022.07.15.500163v1 (2022)","DOI":"10.1101\/2022.07.15.500163"},{"key":"37233_CR85","doi-asserted-by":"publisher","first-page":"5355","DOI":"10.1021\/bi000060h","volume":"39","author":"JL Battiste","year":"2000","unstructured":"Battiste, J. L. & Wagner, G. Utilization of Site-Directed Spin Labeling and High-Resolution Heteronuclear Nuclear Magnetic Resonance for Global Fold Determination of Large Proteins with Limited Nuclear Overhauser Effect Data \u2020. Biochem.-us 39, 5355\u20135365 (2000).","journal-title":"Biochem.-us"}],"container-title":["Nature Communications"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.nature.com\/articles\/s41467-023-37233-1.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.nature.com\/articles\/s41467-023-37233-1","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.nature.com\/articles\/s41467-023-37233-1.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,12,10]],"date-time":"2023-12-10T12:03:39Z","timestamp":1702209819000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.nature.com\/articles\/s41467-023-37233-1"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,4,10]]},"references-count":85,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2023,12]]}},"alternative-id":["37233"],"URL":"http:\/\/dx.doi.org\/10.1038\/s41467-023-37233-1","relation":{},"ISSN":["2041-1723"],"issn-type":[{"value":"2041-1723","type":"electronic"}],"subject":["General Physics and Astronomy","General Biochemistry, Genetics and Molecular Biology","General Chemistry","Multidisciplinary"],"published":{"date-parts":[[2023,4,10]]},"assertion":[{"value":"27 July 2022","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"7 March 2023","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"10 April 2023","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"B.K.K. is a co-founder of and consultant for ConfometRx, Inc. The remaining authors declare no competing interests.","order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Competing interests"}}],"article-number":"2005"}}