Isolation and Characterization of a Novel Angiotensin-Converting Enzyme-Inhibitory Tripeptide from Enzymatic Hydrolysis of Soft-Shelled Turtle (Pelodiscus sinensis) Egg White: In Vitro, In Vivo, and In Silico Study

• Published in: Journal of agricultural and food chemistry 2014-12, Vol.62 (50), p.12178-12185
• Main Authors: Rawendra, Reynetha D. S, Aisha, Chen, Sin-Hong, Chang, Chi-I, Shih, Wen-Ling, Huang, Tzou-Chi, Liao, Ming-Huei, Hsu, Jue-Liang
• Format: Article
• Language: English
• Subjects: active sites; Angiotensin-Converting Enzyme Inhibitors - administration & dosage; Angiotensin-Converting Enzyme Inhibitors - chemistry; Angiotensin-Converting Enzyme Inhibitors - isolation & purification; animal disease models; Animals; Antihypertensive Agents - administration & dosage; Antihypertensive Agents - chemistry; Antihypertensive Agents - isolation & purification; antihypertensive effect; Biocatalysis; Blood Pressure; body weight; digestion; drugs; egg albumen; Egg White - chemistry; eggs; enzymatic hydrolysis; Humans; hypertension; Hypertension - drug therapy; Hypertension - physiopathology; inhibitory concentration 50; Kinetics; Male; Molecular Docking Simulation; oral administration; Pelodiscus sinensis; Peptides - administration & dosage; Peptides - chemistry; Peptides - isolation & purification; peptidyl-dipeptidase A; Peptidyl-Dipeptidase A - chemistry; Peptidyl-Dipeptidase A - metabolism; Rats; Rats, Inbred SHR; systolic blood pressure; trypsin; Trypsin - chemistry; Turtles
• ISSN: 0021-8561; 1520-5118
• DOI: 10.1021/jf504734g

In this study, a novel angiotensin-converting enzyme (ACE)-inhibitory tripeptide (IVR) was isolated and identified from unfertilized soft-shelled turtle egg white (SSTEW). The IC50 value of IVR was measured in vitro as low as 0.81 ± 0.03 μM, and its inhibition type was suggested as competitive according to the Lineweaver–Burk plot. This peptide can be generated from either thermolysin followed by trypsin digestion (two stages) or only trypsin digestion (one stage). Quantitative LC–MS/MS analysis indicated that two-stage digestion gave 3.14 ± 0.17 mg of IVR from 1 g of SSTEW, better than that from one-stage digestion (1.31 ± 0.12 mg). In vivo antihypertensive activity of the tripeptide IVR after single oral administration (0.1 and 1 mg/kg of body weight) led to a significant reduction in systolic blood pressure 2–4 h after administration in spontaneously hypertensive rats. In addition, the binding mechanism of IVR has been rationalized through docking simulations using the testicular ACE (tACE)–lisinopril complex at 2 Å resolution (PDB ). The best docking pose was located at the tACE catalytic site resembling the mode of inhibition exerted by lisinopril, an effective hypertensive synthetic drug. The degree of inhibition of this peptide correlated with the H-bond interaction between the C-terminal of IVR and Lys511 and Tyr520 residues of tACE, a significant inhibitor registration for lisinopril. This study illustrated that IVR behaves as a transition-state analogue inhibitor and is useful in therapeutic intervention for blood pressure control. To the best of our knowledge, this is the first report of an efficient ACE-inhibitory tripeptide generated from the unfertilized egg of soft-shelled turtle.