Heterologous Expression, Antihypertensive Effect, and Underlying Mechanism of Sequence-Designed ACE Inhibitory Peptide YWLKP.

Rational sequence optimization is essential for transforming food-derived peptides into more potent angiotensin-converting enzyme (ACE) inhibitors. LYPVK and its sequence-designed derivative YWLKP exhibit potent activity; however, their binding mechanisms, transport routes, and in vivo antihypertensive effects remain unclear. Here, they were systematically investigated by biotechnological preparation, biolayer interferometry (BLI), molecular dynamics (MD) simulation, Caco-2 transport assays, and studies on spontaneously hypertensive rats (SHRs). BLI revealed that YWLKP bound to ACE with a subnanomolar affinity constant KD of (1.08 ± 0.40) × 10-9 M, consistent with MD results showing a more stable peptide-ACE complex. Caco-2 monolayer transport showed that LYPVK mainly permeated via the paracellular pathway, whereas YWLKP exhibited PepT1-mediated transport. In SHRs, both peptides significantly reduced blood pressure after single and long-term administration, accompanied by renin-angiotensin system modulation, reduced renal and cardiac impairment, and improved endothelial dysfunction. Collectively, these findings support YWLKP as a promising antihypertensive candidate for functional food applications.