Identification of two novel dipeptidyl peptidase-IV inhibitory peptides from sheep whey protein and inhibition mechanism revealed by molecular docking

In this study, the peptides with dipeptidyl peptidase-IV (DPP-IV) inhibitory activities were obtained by trypsin hydrolysis from sheep whey protein. The results showed that the highest DPP-IV inhibitory activity of the hydrolysate (77.80% ± 1.87%) was obtained by trypsin enzymolysis for 4.0 h with 24.50% ± 0.05% of the degree of hydrolysis (DH). Peptides with inhibitory activity against DPP-IV were purified using anion-exchange (DEAE-52) and size-exclusion (G15 dextran gel) chromatography. A total of 86 peptides were identified using LC-MS/MS. The binding energies of RLYLHENK(RL8) and MQEHFTCCR(MQ9) to DPP-IV were determined to be -11.29 kcal/mol and -10.79 kcal/mol with molecular docking in silico, respectively. RL8 and MQ9 could bind to DPP-IV mainly through hydrogen bonds and hydrophobic interactions, and inhibit the DPP-IV enzyme by occupying the S2 pocket. The IC₅₀ values of RL8 and MQ9 in vitro were 166.4 μmol/L and 214.8 μmol/L, respectively. The inhibitory activities in situ modes of the two peptides were evaluated using Caco-2 cells. The IC₅₀ values of RL8 and MQ9 in situ were 158.3 μmol/L and 251.6 μmol/L, respectively. The RL8 and MQ9 derived from sheep whey protein can be considered as a potential source of natural DPP-IV inhibitor.