Identification and molecular mechanism of novel ACE inhibitory peptides from goat milk protein A combined in silico and in vitro study

Fig.1 General description of the proteolytic system of goat milk and the distribution of bioactive peptide species based on online simulation. (A)The enrichment ability of different enzymatic hydrolysis systems to inhibit the function of ACE peptides. (B, C) Distribution of bioactive peptide species after enzymatic hydrolysis of casein and whey proteins.
Fig.2 Study of the screening of conditions and components of goat milk proteolysis. (A-D) Optimal digestion conditions for casein in proteinase K and papain. (E-H) Optimal digestion conditions for whey proteins in proteinase K and papain. (I, K) Effect of enzyme addition sequence on the degree of protein hydrolysis and ACEI rate in goat milk. (J, L) Distribution of ACEI rates for different components, casein and whey proteins.
Fig.3 The 19 potential ACEI peptides were virtually screened from 698 active peptides. (A) The distribution of 698 active peptides. (B) Potential ACEI peptides were virtually screened by PeptideRanker, and CDocker. (C) Molecular weight, logarithmic value of peptide identification score and intensity by LC-MS/MS, and source distribution of enzymatic hydrolysis of 224 potential ACEI peptides. (D) Molecular weight, sequence, logarithmic value of peptide identification score by LC-MS/MS, and source distribution of potential ACEI peptides for 19 of the 698 peptides.
Fig.4 Underlying molecular mechanisms of identified peptides from goat milk against ACE revealed by molecular docking. (A-I) Three-dimensional and two-dimensional diagrams of FKF (A), FRY (B), MPFPK (C), RWL (D), WFK (E), WKP (F), VPP (G), IPP (H), and lisinopril (I) binding to ACE, respectively. (J) The statistics of non-bonded interaction types of ligands (FKF, FRY, MPFPK, RWL, WFK, WKP, VPP, IPP, and lisinopril) interacting with ACE. (K) Heatmap analysis of the number of conventional hydrogen bonds interacting with ACE (S1, S2, S1’, and Zn (II) are the core residues of active sites with ACE). Numerical values indicate the quality and number of conventional hydrogen bonds interactions with active pockets in ACE.
Fig.5 Changes in systolic blood pressure (SBP) in spontaneously hypertensive rats after administration of Saline Solution (A), Captopril (5 mg/kg ip, B) or synthetic peptides (100 mg/kg ip): FKF (C) and FRY (D).