Biochemistry of the Thrombin-Like enzyme and Its Purification from Iranian Echis carinatus snake venom: its interaction with platelet receptors

Document Type : Original Articles

Authors

1 Student Research Committee, Pharmaceutical Sciences Research Centre, School of Pharmacy, Shahid Beheshti University of Medical Sciences, Tehran, Iran.

2 Department of Toxicology, School of Pharmacy, Shahid Beheshti University of Medical Sciences, Tehran, Iran

3 Department of Venomous Animals and Anti-venom, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization, Karaj, Iran

4 Department of Molecular Genetic and Animal Breeding, Gorgan University of Agricultural Sciences and Natural Resources, Golestan, Iran

5 Blood Transfusion Research Center, Institute for Research and Education in Transfusion Medicine, Tehran, Iran

10.32592/ARI.2023.78.6.1822

Abstract

Snake venoms are rich in valuable substances that have medical potential in the diagnosis and treatment of hemostatic diseases. The present paper was aimed at the purification and functional characterization basis of a thrombin-like enzyme and its role in the functioning of the coagulation cascade and platelet aggregation pathway. A thrombin-like serine protease was purified from the Iranian Echis carinatus venom (TLIECV), employing a one-step chromatographic procedure. This peptide was collected in high yield and purity by a single chromatographic step using RP-HPLC equipped with a C18 column. This peptide showed a 3000 Da molecular weight in gel-electrophoresis. Evidence in the SDS-PAGE gel has confirmed high recovery of fraction in optimal terms. Subsequently, this peptide was identified via its intact molecular mass and peptide mass fingerprint (PMF) using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS). Multiple sequence alignments were performed by ClustalW, the Bioedit software. Molegro Data Modeller (MDM) 3.0 software was used to predict the putative tertiary structure of the peptide. The enzyme possessed fibrinogenolytic, procoagulant, and aggregation inducer properties. Moreover, the SDS-PAGE (12%) was applied to examine fibrinogenolytic function. The purified enzyme degraded the Aα chain of fibrinogen while the Bβ and γ chains were not digested. According to that, the deficient human plasma in factor X and normal human plasma were also coagulated by TLIECV, it takes part in the common and intrinsic routes of the coagulation cascade. These findings proved that TLIECV is a serine protease identical to procoagulant thrombin-like snake venom proteases; however, it specifically releases the Aα chain of bovine fibrinogen. Because of its function to make up for the deficiency of factor X and its platelet aggregation inducer property, TLIECV could be considered a molecular impact to reveal the hemostasis mechanisms.

Keywords

Main Subjects

References

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Archives of Razi Institute
Volume 78, Issue 6
November and December 2023
Pages 1822-1835

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