The glucagon-like peptide-1 receptor (GLP-1R) is a class B G protein-coupled receptor (GPCR) central to metabolic regulation, and its potential modulation by dietary phytochemicals is increasingly recognized as physiologically relevant. Understanding how such compounds interact with GLP-1R is important for clarifying mechanisms that may contribute to gut-to-brain signaling. In this study, we examined three structurally related dietary ginsenosides, Rg1, Rg2, and Rg3, as potential modulators of GLP-1R using luciferase reporter assays and computational analyses. Despite sharing similar molecular weights, a common dammarane scaffold, and comparable sugar moieties, the three ginsenosides displayed distinct effects on GLP-1R activity: Rg2 and Rg3 potently reduced receptor activation in a dose-dependent manner when co-administered with Exendin-4, whereas Rg1 had minimal effect. Computational screening of the GLP-1R structure for binding sites identified a putative extracellular pocket on the protein that can accommodate these compounds, while molecular docking and binding free energy calculations provided predicted affinities qualitatively reflecting the phytochemicals’ experimental activities. These findings point to a plausible extracellular mechanism through which dietary ginsenosides may influence GLP-1R responsiveness at the intestinal interface. Our results point to the possibility that non-absorbed phytochemicals can differentially modulate gut-expressed receptors, suggesting a novel pathway for dietary signaling relevant to ethnopharmacology and metabolic health.

The gonadotropins follicle stimulating hormone (FSH) and luteinizing hormone (LH) are key regulators of sexual development and the reproductive cycle in vertebrates. Unlike most G protein-coupled receptors (GPCR), the FSHR and LHR have large extracellular domains containing multiple leucine-rich repeats, which leads to an elaborate mechanism of receptor activation via orthosteric sites that is difficult to manipulate synthetically. To bypass the orthosteric mechanism, in this study using carp as a model organism we identified allosteric sites capable of receptor activation on the transmembrane domain, which are spatially separated from the orthosteric sites. We have further generated pharmacophore hypothesis based on the structural motifs and exposed residues of these cavities. Using available online small compound libraries consisting of >70000 small molecules, we have thereon used receptor cavity-based hypothesis and other screening stages to identify potential modulators of the allosteric binding site on the carp FSHR and LHR in-silico . We then examined by in vitro transactivation assay the effect of four candidate compounds on FSHR and LHR, as compared to the activity of native ligands. Our results reveal both specific and dual effective allosteric modulators for FSHR and LHR, demonstrating the potential of our approach for efficient pharmacophore-based screening.