Identification of bothrojaracin-like proteins in snake venoms from Bothrops species and Lachesis muta.
Bothrojaracin, a 27 kDa protein isolated from Bothrops jararaca venom, forms a non-covalent complex with thrombin, thus blocking its activity. We have previously identified a bothrojaracin-like protein in B. alternatus venom [Castro, H.C., Dutra, D.L.S., Oliveira-Carvalho, A.L., Zingali, R.B., 1998. Bothroalternin, an inhibitor of thrombin from the venom of Bothrops alternatus. Toxicon 36, 1903-1912]. In this report, we have examined snake venoms from six different Bothrops species (B. atrox, B. cotiara, B. jararacussu, B. moojeni and B. neuwiedi), from Lachesis muta and from Crotalus durissus terrificus for the presence of bothrojaracin-like proteins, which we define here as 27 kDa proteins that are immunologically related to bothrojaracin and that inhibit thrombin-induced platelet aggregation. The immunological analysis of these venoms by different techniques indicated the existence of at least one protein recognized by anti-bothrojaracin serum in all venoms tested. Bothrojaracin-like proteins were purified from all crude venoms, except for C. d. terrificus, by a single-step procedure using a thrombin affinity column (PPACK-thrombin-Sepharose). Retained material that inhibits thrombin-induced platelet aggregation was found in a different proportion in each species. Under non-reducing conditions, SDS-PAGE of this material revealed several bands between 20-60 kDa; only those bands corresponding to 27 kDa were recognized by anti-bothrojaracin serum. ELISA confirmed the greater bothrojaracin immunoreactivity of proteins present in B. atrox and B. cotiara as compared to other Bothrops species. Smaller amounts of proteins related to bothrojaracin were found in L. muta venom and were absent from the venom of C. d. terrificus. Our results thus suggest that bothrojaracin-like proteins are widely distributed among Bothrops genera.
Departamento de Bioquímica Médica, ICB/CCS, Universidade Federal do Rio de Janeiro, Brazil.
H C
HC
Castro
M
M
Fernandes
R B
RB
Zingali
D000818:Animals; D017837:Bothrops; D002846:Chromatography, Affinity; D003435:Crotalid Venoms / Q000737:chemistry* / Q000302:isolation & purification / Q000633:toxicity; D004586:Electrophoresis / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay; D005779:Immunodiffusion; D010974:Platelet Aggregation / Q000187:drug effects; D011817:Rabbits; D013045:Species Specificity; D014757:Viper Venoms / Q000737:chemistry*; D017819:Viperidae
D003435:Crotalid Venoms; D014757:Viper Venoms; C083739:bothrojaracin