Exploration of novel dipeptidyl peptidase-IV inhibitory tetrapeptide from walnut (Juglans sigillata) protein and its effect on diabetes-like phenotypes induced by high-sugar diet in Drosophila melanogaster.

Walnut protein was hydrolyzed by different enzymes, and bromelain protein hydrolysate (W-Bromelain) showed the highest dipeptidyl peptidase-IV (DPP-IV) inhibitory activity. W-Bromelain was fractionated, and three novel tetrapeptides (LPQF, LPSF, and VPFP) were identified. In vitro evaluation showed that LPQF exhibited the highest DPP-IV inhibitory activity with an IC value of 99.34 μM. Evaluation of the absorption, distribution, metabolism, excretion, and toxic properties of LPQF showed high absorption and nontoxicity. Molecular docking showed that LPQF could interact with the active residues of DPP-IV through seven hydrogen bonds and five hydrophobic interactions. Molecular dynamics simulation further confirmed the stability of the LPQF-DPP-IV complex. LPQF showed high stability in in vitro gastrointestinal digestion. LPQF ameliorated the type 2 diabetes mellitus-like phenotype and reduced the degree of oxidative stress and intestinal barrier damage in a Drosophila melanogaster model of insulin resistance. Furthermore, the RNA-seq analysis showed that LPQF may exert hypoglycemic effects by regulating the Wnt, MAPK, and FoxO pathways.