Many angiotensin-converting enzyme (ACE) inhibitory peptides (ACEIPs) have been discovered in milk casein hydrolysates, but their inhibitory mechanisms mostly remain unclear. Among these peptides, the tripeptides with proline at the C-terminus are valuable for research due to their stability and absorbability. Analyzing them is time-consuming as no effective screening methods exist. To address this, we constructed a quantitative structure-activity relationship (QSAR) model (R2 = 0.862) and successfully identified a novel ACEIP, Val-Ala-Pro (VAP), from milk casein hydrolysates. Subsequent analysis utilizing isothermal titration calorimetry, multi-spectroscopy, and molecular simulations revealed that VAP is a non-competitive inhibitor with significant ACE activity (IC50 = 1.93 ± 0.17 μM). It binds to key ACE sites (Asn-30, Glu-107, Val-315, Trp-321, and Phe-476) via non-covalent interactions. Moreover, VAP is absorbed in the intestine through paracellular diffusion and the PepT1 pathway. Overall, this work has advanced the research and development of ACEIPs derived from milk casein hydrolysates.