Identification and characterization of WSPSGR: A bioactive peptide from walnut protein hydrolysates with IDO1 inhibitory activity

Previous studies have demonstrated that walnut protein hydrolysates (WPH), obtained through specific enzymatic hydrolysis, exhibit IDO1 inhibitory activity and effectively alleviate depression-like behavior in mice. However, the key bioactive peptide sequences responsible for this effect and their underlying mechanisms remain unclear. In this study, WSPSGR, a bioactive peptide from WPH, was identified as a potent IDO1 inhibitor through virtual screening and experimental validation. Enzymatic and cellular assays demonstrated strong inhibitory activity, with IC 50 values of 0.115 μmol/L and 0.651 μmol/L, respectively. Kinetic analysis confirmed a reversible competitive inhibition mechanism, which was further supported by surface plasmon resonance (SPR) analysis indicating high binding affinity (K D = 1.205 μmol/L). Molecular docking and molecular dynamics (MD) simulations revealed stable interactions at the IDO1 active site, highlighting its structural stability and binding specificity. Furthermore, ADMET analysis suggested good absorption potential and digestive stability, supporting its feasibility for in vivo applications. Simulated gastrointestinal digestion and Caco-2 monolayer permeability studies further confirmed WSPSGR’s stability and intestinal absorption potential. These findings provide new insights into the functional role of walnut-derived peptides in IDO1 inhibition.