Poster 174 - Effects Of recombinant Lopap. Transcriptome and proteome analysis of Lonomia obliqua

Poster 174. Congresso da Sociedade Brasileira de Toxinologia, 8., Symposium of the Pan American Section of the International Society on Toxinology, 8., 2004, Angra dos Reis, Brasil. Abstracts... J. Venom. Anim. Toxins incl.Trop. Dis., 2004, 10, 3, p.533.

Effects Of Recombinant Lopap.Transcriptome And Proteome Analysis Of Lonomia obliqua Bristles.

^{1Reis, C.V.; ¹Flores, M.P.A.; ¹Batista, I.F.C.; ¹Fritzen, M.; ²Junqueira-de-Azevedo, I.L.M; ²Ho, P.L.;} Farsky, ³S.H.P.; ⁴Angles-Cano, E.; ⁵Guillin, M.C.; ¹Chudzinski-Tavassi, A.M.

1 Lab. Bioquímica e Biofísica; 2 Centro de Biotecnologia, I. Butantan, SP; 3 Departamento de Análises Clínicas e Toxicológicas, FCF, USP, SP, Brazil; 4 INSERM U 460; 5 INSERM E9907, Paris, France.

The accidental contact with the Lonomiaobliquacaterpillar venom causes a severe consumptive coagulopathy. In this study, we characterised the active recombinat Lopap (prothrombin activator – 21 kDa) obtained from the cDNA library constructed in pGEM 11 zf(+) plasmid and subcloned in E.coliexpression vector. The r-Lopap activity into the prothrombin activation was assessed in the presence and absence of the prothrombinase complex compounds and the fragments comparison after the prothrombin hydrolysis in different conditions were analysed by SDS-PAGE and by colorimetric substrate hydrolysis. The incubation of r-Lopap with prothrombin showed prethrombin 2 and thrombin generation independently of the prothrombinase complex. According to this we can suggest that the Lopap may be a new type of prothrombin activator. rLopap infusion in mice induces a consumption coagulopathy by fibrinogen depletion (dose dependent manner) similar to that observed with native Lopap. The rLopap prevents the HUVECs apoptosis and increases the expression of NO and PGI2. To cheek out the L. obliqua cDNA and protein profiles, both the transcriptome and proteome were performed. Several clones from the library were sequenced and carried out a comprehensive analysis of gene-expression profiles using ESTs (GenBank) to identify genes involved in the human accident such as toxins involved in physiological system (Lopap, BPP-like, serpins among others). In the same way, the proteome was studied, and its profile showed approximately 120 protein spots, in a pH range between 4 and 9 and MW below 50 kDa. The most representative spots were selected to be analyzed by Mass Spectrometry.