Click
here to close Hello! We notice that
you are using Internet Explorer, which is not supported by Echinobase
and may cause the site to display incorrectly. We suggest using a
current version of Chrome,
FireFox,
or Safari.
Mar Drugs
2024 Feb 16;222:. doi: 10.3390/md22020090.
Show Gene links
Show Anatomy links
Targeted Affinity Purification and Mechanism of Action of Angiotensin-Converting Enzyme (ACE) Inhibitory Peptides from Sea Cucumber Gonads.
Wang Y
,
Chen S
,
Shi W
,
Liu S
,
Chen X
,
Pan N
,
Wang X
,
Su Y
,
Liu Z
.
???displayArticle.abstract???
Protein hydrolysates from sea cucumber (Apostichopus japonicus) gonads are rich in active materials with remarkable angiotensin-converting enzyme (ACE) inhibitory activity. Alcalase was used to hydrolyze sea cucumber gonads, and the hydrolysate was separated by the ultrafiltration membrane to produce a low-molecular-weight peptide component (less than 3 kDa) with good ACE inhibitory activity. The peptide component (less than 3 kDa) was isolated and purified using a combination method of ACE gel affinity chromatography and reverse high-performance liquid chromatography. The purified fractions were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS), and the resulting products were filtered using structure-based virtual screening (SBVS) to obtain 20 peptides. Of those, three noncompetitive inhibitory peptides (DDQIHIF with an IC50 value of 333.5 μmol·L-1, HDWWKER with an IC50 value of 583.6 μmol·L-1, and THDWWKER with an IC50 value of 1291.8 μmol·L-1) were further investigated based on their favorable pharmacochemical properties and ACE inhibitory activity. Molecular docking studies indicated that the three peptides were entirely enclosed within the ACE protein cavity, improving the overall stability of the complex through interaction forces with the ACE active site. The total free binding energies (ΔGtotal) for DDQIHIF, HDWWKER, and THDWWKER were -21.9 Kcal·mol-1, -71.6 Kcal·mol-1, and -69.1 Kcal·mol-1, respectively. Furthermore, a short-term assay of antihypertensive activity in spontaneously hypertensive rats (SHRs) revealed that HDWWKER could significantly decrease the systolic blood pressure (SBP) of SHRs after intravenous administration. The results showed that based on the better antihypertensive activity of the peptide in SHRs, the feasibility of targeted affinity purification and computer-aided drug discovery (CADD) for the efficient screening and preparation of ACE inhibitory peptide was verified, which provided a new idea of modern drug development method for clinical use.
2021FJSC2Y03 Seed Industry Innovation and Industrialization project of Fujian province, FJHJF-L-2022-1 Fujian Province Marine economy development special fund project, FJHJF-L-2022-9 Fujian Province Marine economy development special fund project
Figure 1. ACE inhibitor activity and molecular weight distribution of ultrafiltration hydrolysates: (A) ACE inhibitory activity assay of different proteases hydrolysates; (B) the ACE-inhibiting activity of the ultrafiltration fractions was assessed at various levels of concentration; (C) IC50 values for ACE inhibitory activity of the ultrafiltration fractions; (D) determination of molecular weight distribution of SCGH using gel permeation chromatography (TSK G2000 gel chromatography columns). Different lowercase letters (a–d) on the bars mean that the difference is significant (p < 0.05).
Figure 2. Purification, column separation, and enzyme adsorption of peptides: (A) ACE–Sepharose 4B column affinity adsorption plot; (B) ACE inhibitory activity of different affinity adsorption components; (C) RP-HPLC separation of the targeted affinity eluent (C18 column); (D) ACE inhibitory activity of the purified component by RP-HPLC. Different lowercase letters (a–c) on the bars mean that the difference is significant (p < 0.05).
Figure 3. Inhibition rate and inhibition type: (A) ACE inhibitory activity was measured for each peptide at a concentration of 1 mg/Ml; (B) ACE inhibitory activity of each peptide at different concentrations; (C) IC50 values of ACE inhibitory activity of each peptide; (D) the Lineweaver–Burk plots of the reactions of ACE in the presence of DDQIHIF; (E) the Lineweaver–Burk plots of the reactions of ACE in the presence of HDWWKER; (F) the Lineweaver–Burk plots of the reactions of ACE in the presence of THDWWKER. [S] = hippuryl-l-histidyl-l-leucine concentration; V = velocity of the reaction. Different lowercase letters (a–e) on the bars mean that the difference is significant (p < 0.05).
Figure 4. The 3D and 2D plots of molecular docking of peptides with ACE: (A) the ACE–DDQYHIF complex; (B) the ACE–HDWWKER complex; (C) the ACE–THDWWKER complex. The chart displays different interactions: salt bridge forces (indicated by the orange dotted line), hydrogen bond forces (indicated by the dashed blue line), hydrophobic interaction forces (indicated by the gray dashed line), π–π stacking forces (indicated by the green dashed line), zinc ion forces (indicated by the purple dashed line), and π–cation interactions (indicated by the yellow dashed lines). In the protonation of the receptor, histidine is protonated under different pH conditions and is HID when the hydrogen atom is at the δ-position nitrogen atom and HIE at the ε-position nitrogen atom, i.e., His513, His353, His383, His387, and His410 are transformed to Hie513, Hie353, Hid383, Hid387, and Hie410.
Figure 5. Molecular dynamics (MD) simulation: (A) RMSD curve of ACE–peptide complex system; (B) Rg curve of ACE–peptide complex system; (C) RMSF curve of ACE–DDQYHIF complex system; (D) RMSF curve of ACE–HDWWKER complex system; (E) RMSF curve of ACE–THDWWKER complex system; (F) ACE protein 3D map; (G) diagram of conformational differences between ACE–peptide complex systems and protein monomers. Different color cartoons in the figure represent different systems: the green cartoon is the ACE monomer protein, the blue cartoon is the ACE–peptide complex system, and the arrow points from the ACE monomer protein to the ACE–peptide complex system. The length of the arrow indicates the size of the difference between the two systems.
Figure 6. SBP changes in SHRs after intravenous administration. MC indicates model control (intravenous 0.9% saline). PC indicates positive control (intravenous 5 mg·kg−1 captopril).
