Screening and inhibition mechanism study of novel ACE-inhibitory peptides from silkworm pupa

Numerous natural angiotensin-converting enzyme (ACE) inhibitors (ACEIs) have been extensively studied and applied for the prevention and control of hypertension. This study conducted sequence analysis and computer screening on ACE inhibitory peptides isolated from silkworm pupa, determining their inhibition types through kinetic analysis. By integrating spectral techniques, molecular docking, and molecular dynamics simulations, we investigated the interaction mechanisms between silkworm-derived active peptides and ACE. The results indicated that three novel high-activity ACE inhibitors were identified through computer screening and in vitro validation of DA201-C macroporous resin-purified peptides. MQGFIPE exhibited the strongest inhibitory activity (IC₅₀ = 0.21 ± 0.036 mg/mL), followed by PNDIL (IC₅₀ = 0.569 ± 0.005 mg/mL) and ADNIPIK (IC₅₀ = 0.849 ± 0.001 mg/mL). Lineweaver-Burk plots suggested that MQGFIPE and PNDIL functioned as mixed-type inhibitors, while ADNIPIK acted as a non-competitive inhibitor. Molecular docking simulations revealed that these peptides inhibit ACE through hydrogen bonding formation and hydrophobic interactions with residues. Additionally, spectral studies demonstrated that the peptides MQGFIPE, ADNIPIK and PNDIL effectively quench the intrinsic fluorescence of ACE, alter its conformation and the microenvironment of aromatic amino acid residues, thereby inhibiting its activity. Molecular dynamics simulations confirmed their stable binding to the ACE complex. These findings suggest that silkworm pupa are a significant natural source of ACE-inhibiting peptides, which could serve as functional components in hypertension dietary supplements.