Bitter taste receptors, particularly TAS2R14, play central roles in discerning a wide array of bitter substances, ranging from dietary components to pharmaceutical agents^1,2. TAS2R14 is also widely expressed in extra-gustatory tissues, suggesting its additional roles in diverse physiological processes and potential therapeutic applications³. Here, we present cryo-electron microcopy structures of TAS2R14 in complex with aristolochic acid, flufenamic acid and compound 28.1, coupling with different G protein subtypes. Uniquely, a cholesterol molecule is observed occupying what is typically an orthosteric site in class A GPCRs. The three potent agonists bind, individually, to the intracellular pockets, suggesting a distinct activation mechanism for this receptor. Comprehensive structural analysis, combined with mutagenesis, and molecular dynamic simulations studies, illuminate the receptor’s broad-spectrum ligand recognition and activation via intricate multiple ligand-binding sites. Additionally, our study uncovers the specific coupling modes of TAS2R14 with gustducin and G_i1 proteins. These findings should be instrumental in advancing our knowledge underlying bitter taste perception and its broader implications in sensory biology and drug discovery.

苦味受体，特别是 TAS2R14，在识别从膳食成分到药剂 ^1,2 等多种苦味物质方面发挥着核心作用。 TAS2R14 也在味觉外组织中广泛表达，表明其在多种生理过程和潜在治疗应用中的附加作用 ³ 。在这里，我们展示了 TAS2R14 与马兜铃酸、氟芬那酸和化合物 28.1 复合物的冷冻电子显微镜结构，并与不同的 G 蛋白亚型偶联。独特的是，观察到胆固醇分子占据了 A 类 GPCR 中典型的正位点。三种有效的激动剂分别与细胞内袋结合，表明该受体具有独特的激活机制。全面的结构分析，结合诱变和分子动力学模拟研究，阐明了受体通过复杂的多个配体结合位点的广谱配体识别和激活。此外，我们的研究揭示了 TAS2R14 与 gustducin 和 G _i1 蛋白的特定偶联模式。这些发现应该有助于增进我们对苦味感知的了解及其对感官生物学和药物发现的更广泛影响。