Scorpion venoms are a complex mixture of components. Among them the most important are peptides, which presents the capacity to interact and modulate several ion channel subtypes, including voltage-gated sodium channels (Na_V). Screening the activity of scorpion toxins on different subtypes of Na_V reveals the scope of modulatory activity and, in most cases, low channel selectivity. Until now there are approximately 60 scorpion toxins experimentally assayed on Na_V channels. However, the molecular bases of interaction between scorpion toxins and Na_V channels are not fully elucidated. The activity description of new scorpion toxins is crucial to enhance the predictive strength of the structural–function correlations of these Na_V modulatory molecules. In the present work a new scorpion toxin (Tf1a) was purified from Tityus fasciolatus venom by RP-HPLC, and characterized using electrophysiological experiments on different types of voltage-gated sodium channels. Tf1a was able to modify the normal function of Na_V tested, showing to be a typical β-NaScTx. Tf1a also demonstrated an unusual capability to alter the kinetics of Na_V1.5.

Studies on the interaction sites of peptide toxins and ion channels typically involve site-directed mutations in toxins. However, natural mutant toxins exist among them, offering insights into how the evolutionary process has conserved crucial sequences for activities and molecular target selection. In this study, we present a comparative investigation using electrophysiological approaches and computational analysis between two alpha toxins from evolutionarily close scorpion species of the genus Tityus, namely, Tst3 and Ts3 from T. stigmurus and T. serrulatus, respectively. These toxins exhibit three natural substitutions near the C-terminal region, which is directly involved in the interaction between alpha toxins and Nav channels. Additionally, we characterized the activity of the Tst3 toxin on Nav1.1-Nav1.7 channels. The three natural changes between the toxins did not alter sensitivity to Nav1.4, maintaining similar intensities regarding their ability to alter opening probabilities, delay fast inactivation, and induce persistent currents. Computational analysis demonstrated a preference for the down conformation of VSD4 and a shift in the conformational equilibrium towards this state. This illustrates that the sequence of these toxins retained the necessary information, even with alterations in the interaction site region. Through electrophysiological and computational analyses, screening of the Tst3 toxin on sodium isoform revealed its classification as a classic α-NaTx with a broad spectrum of activity. It effectively delays fast inactivation across all tested isoforms. Structural analysis of molecular energetics at the interface of the VSD4-Tst3 complex further confirmed this effect.

An insect neuroactive helix ring peptide called U₁₁-MYRTX-Tb1a (abbreviated as U₁₁) from the venom of the ant, Tetramorium bicarinatum. U₁₁ is a 34-amino-acid peptide that is claimed to be one of the most paralytic peptides ever reported from ant venoms acting against blowflies and honeybees. The peptide features a compact triangular ring helix structure stabilized by a single disulfide bond, which is a unique three-dimensional scaffold among animal venoms. Pharmacological assays using Drosophila S2 cells have demonstrated that U₁₁ is not cytotoxic but instead suggest that it may modulate potassium channels via the presence of a functional dyad. In our work described here, we have tested this hypothesis by investigating the action of synthetically made U₁₁ on a wide array of voltage-gated K and Na channels since it is well known that these channels play a crucial role in the phenomenon of paralysis. Using the Xenopus laevis oocyte heterologous expression system and voltage clamp, our results have not shown any modulatory effect of 1 μM U₁₁ on the activity of Kv1.1, Kv1.3, Kv1.4, Kv1.5, Shaker IR, Kv4.2, Kv7.1, Kv10.1, Kv11.1 and KQT1, nor on DmNav and BgNav. Instead, 10 μM U₁₁ caused a quick and irreversible cytolytic effect, identical to the cytotoxic effect caused by Apis mellifera venom, which indicates that U₁₁ can act as a pore-forming peptide. Interestingly, the paralytic dose (PD₅₀) on blowflies and honeybees corresponds with the concentration at which U₁₁ displays clear pore-forming activity. In conclusion, our results indicate that the insecticidal and paralytic effects caused by U₁₁ may be explained by the putative pore formation of the peptide.