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The Recombinant Disintegrins Jarastatin and Jararacin and their Inhibitory Activity on Platelet and Endothelial Adhesion

1Universidade Federal do Rio de Janeiro, Instituto de Bioquímica Médica Leopoldo de Meis, Rio de Janeiro, Brazil, 2Universidade do Estado do Rio de Janeiro, Departamento de Biologia Celular, Rio de Janeiro, Brazil, 3Universidade Federal do Rio de Janeiro, Faculdade de Farmácia, Rio de Janeiro, Brazil, 4Universidade Federal do Rio de Janeiro, Instituto de Biofísica Carlos Chagas Filho, Rio de Janeiro, Brazil

Abstract Number: PB1741

Meeting: ISTH 2020 Congress

Theme: Platelets and Megakaryocytes » Platelet Receptors

Background: Integrins are transmembrane heterodimeric glycoproteins, present in almost all cell types, binding extracellular matrix proteins to cytoskeleton. The activation of the integrin by the interaction with its bind promotes the activation of pathways that regulate adhesion, proliferation, differentiation, migration and apoptosis, thus modulating physiological and pathological processes. For this reason, the integrins has been the target of new antithrombotic drugs. The disintegrins from snake venoms are peptides capable of modulating the activity of integrins, among them the αIIbβ3, responsible for the platelet aggregation and αvβ3, related to angiogenesis.

Aims: The aim of this study was to express and analyse the activity toward platelets of recombinant disintegrins jarastatin (rJast) and jararacin (rJarc).

Methods: The secondary structure and their biological activities were analyzed. We performed the expression of these disintegrins in Pichia pastoris, using synthetic gene in the vector pPIC9. They were expressed and secreted in the cultured media and were purified using molecular exclusion chromatography. We confirmed the molecular mass and internal sequence the by mass spectrometry and confirmed protein folding by Circular Dichroism and 1H Nuclear Magnetic Resonance spectra.

Results: The yield of the rJast and rJarc were approximately 40 and 30 mg/ 1L of culture, respectively. rJast and rJarc inhibited platelet aggregation induced by ADP, collagen and thrombin. Both disintegrins inhibited the adhesion of platelets to collagen under continuous flow. We also evaluated effect of rJast on HMEC-1 cells. The viability of the cells was not altered, even with 10 µM of disintegrin. rJast significantly inhibited the adhesion of these cells to vitronectin as well as HMEC-1 migration.

Conclusions: We expressed two RGD disintegrins that showed correct folding and inhibition of platelet aggregation similar to the natives proteins. Finally, these proteins can be used as tools to understand the role of integrins in various physiological and pathological systems including thrombosis and angiogenesis.

To cite this abstract in AMA style:

Zingali R, Succar B, David V, Saldanha-Gama R, Ceneviva Lacerda de Almeida F, Wermelinger Serrão L, Barja-Fidallgo C, Kurtenbach E. The Recombinant Disintegrins Jarastatin and Jararacin and their Inhibitory Activity on Platelet and Endothelial Adhesion [abstract]. Res Pract Thromb Haemost. 2020; 4 (Suppl 1). https://abstracts.isth.org/abstract/the-recombinant-disintegrins-jarastatin-and-jararacin-and-their-inhibitory-activity-on-platelet-and-endothelial-adhesion/. Accessed September 29, 2023.

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