Steviol rebaudiosides bind to four different sites of the human sweet taste receptor (T1R2/T1R3) complex explaining confusing experiments

Sucrose provides both sweetness and energy by binding to both Venus flytrap domains (VFD) of the heterodimeric sweet taste receptor (T1R2/T1R3). In contrast, non-caloric sweeteners such as sucralose and aspartame only bind to one specific domain (VFD2) of T1R2, resulting in high-intensity sweetness. In this study, we investigate the binding mechanism of various steviol glycosides, artificial sweeteners, and a negative allosteric modulator (lactisole) at four distinct binding sites: VFD2, VFD3, transmembrane domain 2 (TMD2), and TMD3 through binding experiments and computational docking studies. Our docking results reveal multiple binding sites for the tested ligands, including the radiolabeled ligands. Our experimental evidence demonstrates that the C20 carboxy terminus of the Gα protein can bind to the intracellular region of either TMD2 or TMD3, altering GPCR affinity to the high-affinity state for steviol glycosides. These findings provide a mechanistic understanding of the structure and function of this heterodimeric sweet taste receptor. Sucrose and other non-caloric sweeteners can bind to different domains of the heterodimeric sweet taste receptor (T1R2/T1R3), resulting in different levels of sweetness. Here, the authors investigate the binding mechanism of various steviol glycosides, artificial sweeteners, and a negative allosteric modulator (lactisole) at four distinct binding sites of T1R2/T1R3 through binding experiments and computational docking studies, revealing multiple binding sites for the tested ligands and structural– function correlations of ligand–receptor interactions.