Upon binding to agonists, G protein-coupled receptors (GPCRs) mediate multiple signaling pathways by coupling to intracellular transducer proteins such as G proteins or -arrestins. These agonists, termed biased ligands, confer functional specificity to GPCRs by activating certain signaling pathways over others. Biased ligands promise precise therapeutic benefits with fewer side effects as drugs compared to today's unbiased GPCR-targeted drugs. Unfortunately due to the paucity of structural data on GPCR-transducer complexes as well as the scarcity of known biased ligands, the molecular mechanisms of biased signaling remain elusive. Obtaining experimental data on the structures of the signaling complexes of GPCRs is daunting since the GPCRs are highly dynamic and technically difficult to isolate and purify in the lab. Consequently, structure-based design of biased ligands for therapeutic and further mechanistic experimental studies has been slow. Progress in understanding the complex signaling landscape of GPCRs can be accelerated if we can increase the success and efficiency of experimental trials. Here, we propose an approach that uses a reliable and time-efficient computational method to guide and accelerate concurrent experiments to stabilize and easily purify GPCR transducer complexes. Such methods need to be developed in tandem with experimental advancements. In the short three-year R01 project our (Vaidehi, Tate and Grisshammer) collaborative efforts have resulted in unprecedented computational methods that markedly increased the understanding of the dynamics of GPCR thermostable mutants and accelerate the purification of GPCRs. The progress we have made in developing and applying novel computational methods has opened up unprecedented opportunities to expand and advance the computational toolbox to identify biasing and thermostabilizing GPCR mutants that can bias the conformations of GPCRs to stably pair with different intracellular transducer proteins, the central process in biased signaling. Building on te successes of the previous R01, we propose to advance our interdisciplinary approach with simultaneous computational method developments and experiments to (1) engineer mutant neurotensin receptor 1 (NTSR1) that shows bias signaling even with unbiased agonist, to study the biased signaling mechanisms of this peptide receptor, (2) advance the computational method LITiConDesign, to predict thermostabilizing mutations for GPCR-transducer complexes, and (3) predict thermostabilizing mutations for avian 1AR-Gs, human A2AR-Gs, 1AR--arrestin1 and A2AR--arrestin1 complexes and verify these predictions with experiments that would provide feedback to improve the computational methods. The outcome of the proposed work is a powerful computational method for routinely predicting biased and thermostable mutants of GPCR-transducer complexes. The method will also accelerate the unraveling of the mechanism of biased signaling in NTSR1 that can be extended easily to other GPCRs.
The much sought-after task of designing 'biased ligands' that reduce the side effects of drugs, remains a challenging one due to the lack of understanding of biased signaling mechanisms in G-protein coupled receptors (GPCRs). The proposed interdisciplinary research will develop computational methods concurrent with experiments to speed up the design of biased and thermo-stabilized GPCR mutants that will stabilize the GPCR- G-protein and GPCR-arrestin complexes. Further purification of these complexes will lead to studies on understanding biased signaling mechanisms and pave way to designing biased ligands for GPCRs for treatment of hypertension, diabetes and cancer.
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| Suno, Ryoji; Lee, Sangbae; Maeda, Shoji et al. (2018) Structural insights into the subtype-selective antagonist binding to the M2 muscarinic receptor. Nat Chem Biol 14:1150-1158 |
| Ghosh, Soumadwip; Bierig, Tobias; Lee, Sangbae et al. (2018) Engineering Salt Bridge Networks between Transmembrane Helices Confers Thermostability in G-Protein-Coupled Receptors. J Chem Theory Comput : |
| Choy, Cecilia; Ansari, Khairul I; Neman, Josh et al. (2017) Cooperation of neurotrophin receptor TrkB and Her2 in breast cancer cells facilitates brain metastases. Breast Cancer Res 19:51 |
| Lee, Sangbae; Mao, Allen; Bhattacharya, Supriyo et al. (2016) How Do Short Chain Nonionic Detergents Destabilize G-Protein-Coupled Receptors? J Am Chem Soc 138:15425-15433 |
| Vaidehi, Nagarajan; Bhattacharya, Supriyo (2016) Allosteric communication pipelines in G-protein-coupled receptors. Curr Opin Pharmacol 30:76-83 |
| Vaidehi, Nagarajan; Grisshammer, Reinhard; Tate, Christopher G (2016) How Can Mutations Thermostabilize G-Protein-Coupled Receptors? Trends Pharmacol Sci 37:37-46 |
| Krumm, Brian E; Lee, Sangbae; Bhattacharya, Supriyo et al. (2016) Structure and dynamics of a constitutively active neurotensin receptor. Sci Rep 6:38564 |
| Bhattacharya, Supriyo; Salomon-Ferrer, Romelia; Lee, Sangbae et al. (2016) Conserved Mechanism of Conformational Stability and Dynamics in G-Protein-Coupled Receptors. J Chem Theory Comput 12:5575-5584 |
| Semack, Ansley; Sandhu, Manbir; Malik, Rabia U et al. (2016) Structural Elements in the G?s and G?q C Termini That Mediate Selective G Protein-coupled Receptor (GPCR) Signaling. J Biol Chem 291:17929-40 |
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