XXXV Reunião Anual da SBBqResumoID:8601


Analysis of Bothrops jararaca coagulation inhibitor (BjI) similar proteins in Bothrops alternatus, Bothrops jararacussu and Crotalus durissus terrificus snake plasmas


Morais KB1; Grego KF2 and Tanaka-Azevedo AM1.



1Laboratório de Fisiopatologia, Instituto Butantan, São Paulo, Brasil; 2Laboratório de Herpetologia, Instituto Butantan, São Paulo, Brasil.


Bothrops jararaca coagulation inhibitor (BjI) is a high molecular weight protein isolated from B. jararaca snake blood which inhibits the thrombin coagulant activity (Tanaka-Azevedo et. al. 2004). The aim of this work was the comparison of anticoagulant effect of Bothrops alternatus, Bothrops jararacussu and Crotalus durissus terrificus snake plasmas and also investigate the presence of BjI similar proteins in these species. The snake blood was collected by caudal punction and the plasma was separated by centrifugation. The same as B. jararaca plasma, the poisonous snake plasmas also presented a prolonged thrombin time (31.6 s, 44.1 s and 45.4 s for B. alternatus, B. jararacussu and C. durissus terrificus, respectively) when compared to control (10 s). The snake plasma proteins  analyzed by SDS-PAGE showed similar profiles. Antibody against BjI was obtained in mice by subcutaneous injection of purified BjI (5 mg) emulsified with 200 mL complete Freund’s adjuvant. The immunization procedure was followed by boosters using the incomplete Freund’s adjuvant. The pre-immune mice serum was used as control for immunological assays. Serum titers were determined by ELISA method. The antiserum containing anti-BjI was used in order to confirm if the prolonged thrombin times found in these snakes plasmas were caused by the presence of protein immunological similar to BjI by western blotting experiments. Our present results confirmed the presence of BjI similar proteins in B. alternatus, B. jararacussu and C. durissus terrificus plasmas by western blotting. The perspective of this study is to purify these proteins in order to compare them with  BjI.

 

Supported by FAPESP (nº 04/02224-4 e 04/13434-0).