Study On Identification And Mechanism Of Angiotensin Ⅰ-Converting Enzyme (ACE) Inhibitory Peptide From Silkworm Protein

With the advantages of safe, non-toxic and so on, angiotensin Ⅰ-converting enzyme (ACE) inhibitory peptide from natural source has become one of the potential alternatives of antihypertensive drug and health care products. Basing on two ACE inhibitory peptides (P1 and P2), in this research, the amino acid sequences of P1 and P2 were determined by MALDI TOF MS; the inhibition mechanisms were studied by inhibition kinetic and molecular docking, and a series of modified peptide with high ACE inhibitory activity were obtained. The main research results are as follows:(1) The amino acid sequences of P1, P2 were determinted by MALDI TOF MS. The amino acid sequence of P1 was Gly-Asn-Pro-Trp-Met (GNPWM) with a molecular weight of 603.76 u, and the amino acid sequence of P2 was Asn-Arg-Tyr-Leu-Arg (NRYLR) with a molecular weight of 720.88 u. Their IC50 were 12.61 μg/mL and 14.68 μg/mL, respectively. Their resistance to high processing temperatures and simulated gastrointestinal digestion environment were studied. P1 had good thermal stability under the condition of 40℃, and could resist simulated digestion; P1 had good thermal stability under the condition of 40℃, but it could not resist simulated digestion.(2) The inhibition mechanisms of PI, P2 were studied. The inhibition of P1, P2 on ACE were reversible inhibition, which were studied by inhibition kinetic; PI was a noncompetitive inhibitor, and the inhibition constant Ki,1=0.037 mmol/L; P2 was a competitive inhibitor, and the inhibition constant Ki,2=0.020 mmol/L. The molecular docking was performed by Sybyl. P1 could form hydrogen bond with the amino acids of ACE Thr166、Gln281、Thr372、Asp377、 Lys511、His513、Tyr520, and P2 could form hydrogen bond with the amino acids of ACE Gln281、Ala356、Glu403、Asp415、Lys5111、Tyr520.(3) A series of peptide were designed by basing on P1 and P2 to obtained peptide with high ACE inhibitory activity. The effects of C terminal amino acid and N terminao amino acid on the ACE inhibitory activity were studied, respectively. The peptides with high ACE inhibitory activity, such as Gly-Asn-Pro-Trp-Trp (IC50=11.0 μg/mL), Asn-Pro-Trp-Trp (IC50=11.9 μg/mL), Trp-Phe (IC50=8.6 μg/mL), Trp-Trp (IC50=4.2 μg/mL), Arg-Tyr-Leu-Met (TC50=8.5 μg/mL), Arg-Tyr-Leu-Arg (IC50=8.2 μg/mL), Tyr-Leu-Met (IC50=7.9 μg/mL), were obtained.