Variability of sodium channel toxins in Mesobuthus thersites Advised toolkit for comprehensive venomics

Natural venoms constitute vast combinatorial libraries of ion channel ligands with unprecedented properties including high affinity and selectivity. Scorpion venoms in particular are abundant in sodium and potassium channel ligands representing small proteins or peptides, which are traditionally studied by transcriptomics and proteomics approaches. However, a universal algorithm to pursue the examination of multicomponent peptide-rich mixtures such as venoms is lacking. Here we present a general scheme to the analysis of scorpion toxin arsenal with focus on sodium channel ligands. We explored the venom of the Central Asian scorpion Mesobuthus thersites using venom gland transcriptomics and three proteomic methods. We compared traditional automated annotation by Blast2GO with our custom approach based on regular expressions. We found that the traditional annotation is compromised yielding a considerable percentage of false annotations, whereas our approach is rigorous. As a result, we predicted 120 unique toxin-like peptides from transcriptomic data, 30 of which were omitted by Blast2GO. Using a combination of MALDI and LC-MS proteomic techniques, we detected the presence of 358 peptides in the venom. Our results demonstrate the limitations of the currently used methods in venomics and suggest a preferred framework for the investigation of toxin variability. Implementation of this framework will aid toxinologists to uncover previously elusive or undetected venom components.