Research

Ontogenetic variations in the venom proteome of the Amazonian snake Bothrops atrox

Rafael AP Guércio¹ , Anna Shevchenko² , Andrej Shevchenko² , Jorge L López-Lozano³ , Jaime Paba¹ , Marcelo V Sousa¹ and Carlos AO Ricart¹

¹  Brazilian Center for Protein Research, Department of Cell Biology, University of Brasilia, Brasília, 70910-900- DF, Brazil

²  Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01307 Dresden, Germany

³  Gerência de Animais Peçonhentos – Fundação de Medicina Tropical do Amazonas, Av. Pedro Teixeira 25, 69040-000 Manaus, AM, Brazil

author email corresponding author email

Proteome Science 2006, 4:11doi:10.1186/1477-5956-4-11

Published:	11 May 2006

Abstract

Background

Bothrops atrox is responsible for the majority of snakebite accidents in the Brazilian Amazon region. Previous studies have demonstrated that the biological and pharmacological activities of B. atrox venom alter with the age of the animal. Here, we present a comparative proteome analysis of B. atrox venom collected from specimens of three different stages of maturation: juveniles, sub-adults and adults.

Results

Optimized conditions for two-dimensional gel electrophoresis (2-DE) of pooled venom samples were achieved using immobilized pH gradient (IPG) gels of non-linear 3–10 pH range during the isoelectric focusing step and 10–20% gradient polyacrylamide gels in the second dimension. Software-assisted analysis of the 2-DE gels images demonstrated differences in the number and intensity of spots in juvenile, sub-adult and adult venoms. Although peptide mass fingerprinting (PMF) failed to identify even a minor fraction of spots, it allowed us to group spots that displayed similar peptide maps. The spots were subjected to a combination of tandem mass spectrometry and Mascot and MS BLAST database searches that identified several classes of proteins, including metalloproteinases, serine proteinases, lectins, phospholipases A₂, L-amino oxidases, nerve growth factors, vascular endothelial growth factors and cysteine-rich secretory proteins.

Conclusion

The analysis of B. atrox samples from specimens of different ages by 2-DE and mass spectrometry suggested that venom proteome alters upon ontogenetic development. We identified stage specific and differentially expressed polypeptides that may be responsible for the activities of the venom in each developmental stage. The results provide insight into the molecular basis of the relation between symptomatology of snakebite accidents in humans and the venom composition. Our findings underscore the importance of the use of venoms from individual specimen at various stages of maturation for the production of antivenoms.