Related Experiment Video
Updated: Jul 29, 2026
Using a GFP-tagged TMEM184A Construct for Confirmation of Heparin Receptor Identity
Published on: February 17, 2017
Identification, screening, and comprehensive evaluation of novel thrombin inhibitory peptides from the hirudo
Xiaoyu Chai1, Fulu Pan1, Qianqian Wang1
1Department of Chemistry of Traditional Chinese Medicine, School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing, China.
Insights
Hirudo hydrolysates (HHS) show promising thrombin inhibitory potential, with a specific peptide (P1) demonstrating significant anticoagulant effects. Further optimization of P1 is recommended for antithrombotic drug development.
Area of Science:
- Biochemistry
- Pharmacology
- Medicinal Chemistry
Background:
- Thrombin inhibition is a key strategy for managing cardiovascular disease (CVD).
- Oral peptide and protein drug efficacy is limited by gastrointestinal digestion.
- Hirudo extracts offer potential as sources of antithrombotic agents.
Purpose of the Study:
- To evaluate the thrombin inhibitory properties of pepsin-produced hirudo hydrolysates (HHS).
- To develop a comprehensive screening and evaluation approach for thrombin inhibitors.
- To identify and characterize potent thrombin inhibitory peptides (TIPs) from HHS.
Main Methods:
- In vitro evaluation of hirudo extract and HHS thrombin inhibitory activity.
- Nano LC-MS/MS coupled with in silico analysis for TIP screening.
- In vitro anticoagulant assays (APTT, TT, PT) and platelet aggregation inhibition.
- UV-Vis spectroscopy and molecular dynamics simulations to study TIP-thrombin interactions.
Main Results:
- HHS retained 60-75% of thrombin inhibition activity.
- 90 peptides were identified in HHS; Asn-Asp-Leu-Trp-Asp-Gln-Gly-Leu-Val-Ser-Gln-Asp-Leu (P1) was the most potent TIP.
- P1 exhibited significant thrombin inhibition (IC50: 2,425.5 ± 109.7 μM), dose-dependently prolonged thrombin time, and reduced platelet aggregation.
- Spectroscopic and simulation data confirmed P1 binding to thrombin.
Conclusions:
- HHS represent a valuable source for discovering and evaluating antithrombotic compounds.
- The identified peptide P1 shows potential for structural optimization and further preclinical evaluation.
- This study provides a robust framework for screening and validating novel thrombin inhibitors.
Purpose:
The inhibition of thrombin has proven to be an efficacious therapeutic approach for managing cardiovascular disease (CVD), with widespread implementation in clinical settings. Oral ingestion of peptides and protein drugs is influenced by gastrointestinal digestive enzymes. We aimed to evaluate the thrombin inhibitory properties of hirudo hydrolysates (HHS) produced by pepsin and propose a comprehensive approach to screen and evaluate thrombin inhibitors.
Methods:
We evaluated the in vitro inhibitory properties of the hirudo extract, both before and after hydrolysis with pepsin, toward thrombin. We screened for the most potent thrombin inhibitory peptide (TIP) using nano liquid chromatography-tandem mass spectrometry (Nano LC-MS/MS) coupled with in silico analysis. Next, we employed the thrombin inhibition activity IC50 to investigate the interaction between TIP and thrombin, and conducted in vitro evaluations of its anticoagulant effects (APTT, TT, PT), as well as its ability to inhibit platelet aggregation. Furthermore, we utilized UV-Vis spectroscopy to explore structural changes in thrombin upon binding with TIP and employed molecular dynamics simulations to delve deeper into the potential atomic-level interaction modes between thrombin and TIP.
Results:
The retention rate of thrombin inhibition for HHS was found to be between 60% and 75%. A total of 90 peptides from the HHS were identified using LC-MS/MS combined with de novo sequencing. Asn-Asp-Leu-Trp-Asp-Gln-Gly-Leu-Val-Ser-Gln-Asp-Leu (NDLWDQGLVSQDL, P1) was identified as the most potent thrombin inhibitory peptide after in silico screening (molecular docking and ADMET). Then, the in vitro study revealed that P1 had a high inhibitory effect on thrombin (IC50: 2,425.5 ± 109.7 μM). P1 exhibited a dose-dependent prolongation of the thrombin time (TT) and a reduction in platelet aggregation rate. Both UV-Vis spectroscopy and molecular dynamics simulations demonstrated that P1 binds effectively to thrombin.
Conclusion:
Overall, the results suggested that HHS provides new insights for searching and evaluating potential antithrombotic compounds. The obtained P1 can be structurally optimized for in-depth evaluation in animal and cellular experiments.
Related Concept Videos
01:30Anticoagulant Drugs: Low-Molecular-Weight Heparins
01:20Blood Studies for Cardiovascular System I: Cardiac Biomarkers
The essential diagnostic tools for detecting myocardial necrosis and monitoring individuals suspected of having acute coronary syndrome (ACS) include:
Troponins
Troponins, particularly cardiac troponins I and T, are the most precise and sensitive markers of myocardial injury. They are detectable within 4-6 hours of myocardial injury and remain...
01:30Dysrhythmias V: Evaluating Dysrhythmias