Unlocking TAS2R14 activation through intricate multi-ligand binding networks

Bitter taste receptors, particularly TAS2R14, play central roles in discerning a wide array of bitter substances, ranging from dietary components to pharmaceutical agents[1][1]–[3][2]. In addition, TAS2R14 has broad expression in non-gustatory tissues, suggesting its important roles in selective physiological processes and therapeutic potential[4][3]. Here, we present cryo-electron microcopy structures of TAS2R14 in complex with flufenamic acid and aristolochic acid, coupling with G protein subtypes gustducin and Gi. Distinct from most known GPCRs, agonists of TAS2R14 bind to multiple intracellular pockets. We highlight the cholesterol molecules occupying an upper transmembrane site, typically the orthosteric pocket in other GPCRs, also binding in the entrance to an intracellular agonist binding pocket, suggesting an endogenous modulatory function[5][4]. The structural and mutagenesis analysis illuminate the receptor’s broad-spectrum ligand recognition and activation via intricate multiple ligand-binding sites. Furthermore, we unveil the structural configuration of gustducin and its interaction with TAS2R14, as well as the coupling mode of Gi1. This investigation should be instrumental in advancing our knowledge of the mechanisms underlying bitter taste recognition and response, with broader relevance to sensory biology and pharmacology. ### Competing Interest Statement The authors have declared no competing interest. The atomic coordinates for AA150µM-TAS2R14-miniGs/gust, AA50µM-TAS2R14-Gi and AA150µM-TAS2R14-Gi have been deposited in the Protein Data Bank with the accession codes 8XQL, 8XQN and 8XQO, respectively. The EM maps for AA150µM-TAS2R14-miniGs/gust, AA50µM-TAS2R14-Gi and AA150µM-TAS2R14-Gi have been deposited in EMDB with the codes EMD-38580, EMD-38582 and EMD-38583, respectively. The atomic coordinates for FFA1mM-TAS2R14-miniGs/gust, and FFA150µM-TAS2R14-Gi, have been deposited in the Protein Data Bank with the accession codes 8XQR and 8XQS, respectively. The EM maps for FFA1mM-TAS2R14-miniGs/gust, and FFA150µM-TAS2R14-Gi, have been deposited in EMDB with the codes EMD-38586, and EMD-38587, respectively. The atomic coordinate for AA150µM-TAS2R14-Gg and Apo-TAS2R14-Gi have been deposited in the Protein Data Bank with the accession codes 8XQP and 8XQT. The EM map for AA150µM-TAS2R14-Gg and Apo-TAS2R14-Gi, have been deposited in EMDB with the code EMD-38584 and EMD-8XQT, respectively. All other data are available upon request to the corresponding authors T.H., Z-J.L. and J.-P.S. [1]: #ref-1 [2]: #ref-3 [3]: #ref-4 [4]: #ref-5