Novel Dual DPP-IV/ACE Inhibitory Peptides from Tartary Buckwheat: From Inhibition Kinetics to Cellular Bioactivity

Type 2 diabetes and hypertension frequently coexist, necessitating multi-target interventions. This study used a virtual-to-experimental pipeline to identify dual dipeptidyl peptidase-IV (DPP-IV)/angiotensin-converting enzyme (ACE) inhibitory peptides from Tartary buckwheat (Fagopyrum tataricum), identifying VPIY and IPLIGF. VPIY showed IC50 (half-maximal inhibitory concentration) values of 191.61 ± 4.99 μM for DPP-IV and 140.85 ± 4.35 μM for ACE, whereas IPLIGF showed 291.43 ± 4.29 μM and 199.85 ± 5.59 μM for DPP-IV and ACE, respectively. Kinetic analyses using global nonlinear mixed-model fitting revealed that VPIY acted as a mixed-type inhibitor of DPP-IV and a non-competitive inhibitor of ACE, whereas IPLIGF was non-competitive against DPP-IV and competitive-like against ACE. Molecular docking confirmed VPIY as a peripheral, Zn2+-non-coordinating ACE inhibitor, whereas IPLIGF ligated the ACE Zn2+, supporting competitive inhibition. Notably, VPIY displayed ~5-fold enhancement over its enzymatic IC50 potency in Caco-2 (human colorectal adenocarcinoma) cells (IC50 = 38.03 ± 2.23 μM). Both peptides restored nitric oxide (NO) and suppressed endothelin-1 (ET-1) in norepinephrine-injured human umbilical vein endothelial cells (HUVECs), mitigating endothelial dysfunction. These first-reported dual-inhibitory peptides from Tartary buckwheat, particularly VPIY, exert potent in situ DPP-IV inhibitory activity on Caco-2 cells and vascular protection, supporting their development as food ingredients for metabolic syndrome.