Receptor-Based Functional Characterization Reveals Synergistic Umami Enhancement of Anchovy Fish Sauce-Derived Peptides with Monosodium Glutamate

Abstract The mechanistic quantification of umami synergy has long been impeded by conventional sensory evaluation and electronic tongues, which fail to dissect multiligand synergistic effects from receptor recognition and signal transduction perspectives. This study established a multidimensional screening strategy integrating machine learning, molecular docking, and cellular validation, identifying four umami peptides (SDHE, NE, TRE, TMHE) from anchovy fish sauce. Sensory tests showed that these peptides had weak umami intensity alone (score 5–6), but significantly enhanced umami intensity (score 6–8) and prolonged aftertaste (80–110 s) when coadded with MSG. Calcium signaling assays using HEK-293 cells stably expressing human T1R1/T1R3 confirmed these peptides act as positive allosteric modulators, amplifying MSG-triggered signals by 1.07–2.29-fold. Molecular docking revealed that cobinding of these peptides with MSG at the Venus flytrap domain increased the number of noncovalent interactions from 5 to 8–17. This study provides direct receptor-level evidence of umami synergy and lays a foundation for discovering natural synergistic umami enhancers.