Isolation and Functional Identification of an Antiplatelet RGD-Containing Disintegrin from Cerastes cerastes Venom
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Isolation and Functional Identification of an Antiplatelet RGD‑Containing Disintegrin from Cerastes cerastes Venom Meriem Ameziani1 · Fatah Chérifi1 · Hamida Kiheli1 · Samah Saoud1 · Ghania Hariti2 · Safia Kellou‑Taîri3 · Fatima Laraba‑Djebari1 Accepted: 17 September 2020 © Springer Science+Business Media, LLC, part of Springer Nature 2020
Abstract The current report focuses on purification, structural and functional characterization of Cerastategrin from Cerastes cerastes venom, a novel basic disintegrin (pI 8.36) with 128 amino acid residues and a molecular weight of 13 835.25 Da measured by MALDI-MSMS. The 3D structure of Cerastategrin is organized as α-helix (13%), β-strand (15%) and disordered structure (30%) and presents homologies with several snake venom disintegrins. Structural modeling shows that Cerastategrin presents an RGD motif that connects specifically to integrin receptors. Cerastategrin exhibits the inhibition of ADP induced platelets with an I C50 of 0.88 µg/mL and shows in vivo long stable anticoagulation effect 24 h post-injection of increasing doses ranging from 0.2 to 1 mg/kg, therefore, Cerastategrin maintained irreversibly the blood incoagulable. Moreover, Cerastategrin decreases the amount of bounded αIIbβ3 and reduced significantly the quantity of externalized P-Selectin. Cerastategrin acts as a molecule targeting specifically the receptor αIIbβ3; therefore, it behaves as a potent platelet activation inhibitor. As a new peptide with promising pharmacological properties, Cerastategrin could have a potential therapeutical effect in the vascular pathologies and may be a new effective treatment approach. Keywords Cerastategrin · Disintegrin · Antiplatelet activity · GPIIbIIIa inhibitor · RGD binding motif Abbreviations Cc1-PLA2 Cerastes cerastes Phospholipase A2 CCSV-MPase Cerastes cerastes Snake venom metalloproteinase SDS PAGE Sodium dodecyl sulphate H2SO4 Sulfuric acid NaOH Sodium hydroxid TFA Trifluorophosphate acide ACN Acetonetrile TCEP Tris(2-carboxyethyl) phosphine IAM 10 MM 2-iodoacetamide * Fatima Laraba‑Djebari [email protected]; [email protected] 1
USTHB, Laboratory of Cellular and Molecular Biology, Faculty of Biological Sciences, BP 32 El‑Alia, Bab Ezzouar, Algiers, Algeria
2
Faculty of Medicine, Blood Transfusion Center, Unverisity of Benyoucef Benkheda Algiers 1, CHU Bab El‑Oued, Algiers, Algeria
3
USTHB, Laboratory of Theoretical Physico-Chemistry and Computer Chemistry, Faculty of Chemistry, BP 32 El‑Alia, Bab Ezzouar, Algiers, Algeria
EDTA Ethylenediaminetetraacetic EGTA Aminopolycarboxylic acid TRAP-6 Thrombin receptor activator peptide-6
1 Introduction Platelets have a crucial role in the hemostasis and thrombosis by adhering to, spreading over subendothelial surfaces, and aggregating to each other [1]. Platelet-platelet bridges are formed and occurred by the binding of fibrinogen to its platelet receptor GPIIbIIIa commonly known as αIIbβ3 integrin. This receptor is a heterodimeric glycoprotein that mediates cell–cell and cell–matrix i
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