Disintegrin activity of Recombinant Insularin and Fractions Isolated from Bothrops jararaca venom

Della-Casa M.S¹.; Tanjoni, I¹.; Junqueira de Azevedo, I.L.M².; Ho, P.L².; and Moura-da-Silva, A. M.¹.

1 Laboratório de Imunopatologia e 2 Centro de Biotecnologia, Instituto Butantan

Disintegrins represent a family of cysteine-rich, low molecular weight proteins occurring in venoms of various vipers. They bind with a high affinity to numerous integrins and are potent inhibitors of platelet aggregation and cell adhesion. We are studying the role of insularin, a recombinant disintegrin from the venom of Bothrop insularis and fractions isolated from the venom of B. jararaca able to inhibit platelet-aggregation. Insularin inhibited ADP-induced platelet aggregation with IC50 of 0,600mM. From B. jararaca venom, we have isolated 23 fractions by HPLC C18-reversed phase column. When these fractions were screened in protein excess (80mg/ml), only fractions 3, 4 and 5 inhibited 100% ADP-induced platelet-aggregation. Further dose-response experiments of those three fractions showed that fraction 3 was the most potent inhibitor of platelet aggregation followed by fraction 5 and 4. Insularin and fractions 3, 4 and 5 were also tested for direct adhesion of culture endothelial cells (tend). Endothelial cells adhered to plates coated with the different disintegrins. In this test, fractions 3 and 5 were the most efficient for the binding, followed by insularin and fraction 4. Our study contributes for the discovery of news antagonists of integrins involved in pathologies dependent on cell adhesion.

Supported by: FAPESP, CNPq and Fundação Butantan. 

Correspondence to: belleclues@yahoo.com