In vitro inhibition of α-amylase and angiotensin converting enzyme by fava bean (Vicia faba) protein-derived peptides, as revealed by kinetics, molecular docking and molecular dynamics

The present study investigated the inhibitory effects of a fava bean ( Vicia faba ) protein-derived peptides on α-amylase and angiotensin-converting enzyme (ACE) using an integrated approach that combined enzyme kinetics, fluorescence spectroscopy, molecular docking, and molecular dynamics (MD) simulation. Pepsin fava bean protein hydrolysate was fractionated by successive RP-HPLC, yielding bioactive fractions enriched in antioxidant and enzyme-inhibitory activities. LC–MS/MS analysis identified seven novel peptides, among which TPACSLPL showed the strongest multifunctional activity. TPACSLPL exhibited concentration-dependent antioxidant activity and significantly inhibited α-amylase (IC 50 = 0.41 mg/mL) and ACE (IC 50 = 0.37 mg/mL) at 1.0 mg/mL. Kinetic analysis revealed competitive inhibition for both α-amylase and ACE, characterized by an increased Km with minimal change in Vmax, indicating that the peptide primarily interferes with the substrate binding at the active site without affecting catalytic turnover. Intrinsic fluorescence spectroscopy confirmed peptide–enzyme interactions, showing fluorescence enhancement in α-amylase and marked quenching in ACE, indicative of stable complex formation. Docking studies demonstrated strong binding affinities (ΔG − 9.8 and −10.1 kcal/mol for α-amylase and ACE, respectively), while 100 ns MD simulations and molecular mechanics with generalized born surface area analysis confirmed structural stability and favourable binding energetics, particularly for ACE. These findings highlight TPACSLPL as a novel, promising multifunctional peptide candidate for managing metabolic syndrome through targeted enzyme inhibition mechanisms.