Mining of Novel Human DPP-IV-Targeting Tripeptides through Multicomputational Screening and In Vitro Validation for...

Inhibition of human dipeptidyl peptidase IV (hDPP-IV) is considered an effective strategy in the management of type 2 diabetes (T2D). However, the lack of high-throughput screening approaches and the high cost of experimental evaluation have limited the discovery of peptide-based hDPP-IV inhibitors. Herein, an integrated multicomputational screening strategy was employed to successfully mine novel hDPP-IV inhibitory tripeptides. Among the screened candidates, FAW exhibited the highest inhibitory activity, with an IC₅₀ of 233 μM, establishing it as a promising scaffold for further development. Structural-based density functional theory (DFT) analysis revealed that active tripeptides exhibit localized electronic regions with higher electrophilicity indices than less active tripeptides. Furthermore, experimental evaluation demonstrated that stereospecific recognition of l-alanine by hDPP-IV plays a critical role in FAW's inhibitory activity. Molecular dynamics simulation further indicated that the FAW-hDPP-IV complex induces less conformational perturbation in the enzyme than the d-alanine-containing analogue. Collectively, the agreement between the multicomputational screening and the in vitro validation results offers a valuable strategy for mining other short-chain bioactive peptide candidates that may inhibit hDPP-IV.