State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China.
State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China; Biodrug Research Center, Wuhan University, Wuhan 430072, China.
Peptides. 2018 Jan;99:153-160. doi: 10.1016/j.peptides.2017.09.017. Epub 2017 Oct 3.
Scorpion toxins are invaluable pharmacological tools for studying ion channels and potential drugs for channelopathies. The long-chain toxins from scorpion venom with four disulfide bridges exhibit their unusual bioactivity or biotoxicity by acting on the sodium channels. However, the functional properties of most toxins are still unclear due to their tiny amounts in crude venom and their challenging production by chemical and gene engineering techniques. Here, we expressed one of the long-chain α-toxins, BmKM9, found in the venom of the scorpion Buthus martensii Karsch and characterized its pharmacological properties on sodium channels. Unlike previous toxin production, the recombinant BmKM9 (rBmKM9) possessed no additional amino acid residues such as the His-tag and thrombin cleavage site. The refolded toxin could inhibit the inactivation of rNa1.4, hNa1.5 and hNa1.7 sodium channels. Dose-response experiments were further conducted on these channels. The calculated EC values were 131.7±6.6nM for rNa1.4, 454.2±50.1nM for hNa1.5 and 30.9±10.3μM for hNa1.7. The channel activation experiments indicated that the rBmKM9 toxin could shift the activation curves of rNa1.4 and hNa1.5 channels toward a more negative direction and present the typical features of a β-toxin. However, instead of the same activation property on sodium channels, the rBmKM9 toxin could result in different inactivation shift capabilities on rNa1.4 and hNa1.5 channels. The V values of the steady-state inactivation were altered to be more positive for rNa1.4 and more negative for hNa1.5. Moreover, the recovery of the hNa1.5 channel from inactivation was more significantly delayed than that of the rNa1.4 channel by exposure to rBmKM9. Together, these findings highlighted that the rBmKM9 toxin presents the pharmacological properties of both α- and β-toxins, which would increase the challenge to the classical classification of scorpion toxins. Furthermore, the expression method and functional information on sodium channels would promote the potential application of toxins and contribute to further channel structural and functional studies.
蝎毒素是研究离子通道的宝贵药理学工具，也是潜在的通道病药物。具有四个二硫键的长链毒素来自蝎毒液，通过作用于钠通道表现出其异常的生物活性或生物毒性。然而，由于粗毒液中蝎毒素的含量非常少，并且通过化学和基因工程技术进行生产具有挑战性，因此大多数毒素的功能特性仍不清楚。在这里，我们表达了一种长链α-毒素 BmKM9，该毒素存在于蝎子 Buthus martensii Karsch 的毒液中，并对其在钠通道上的药理学特性进行了表征。与之前的毒素生产方法不同，重组 BmKM9（rBmKM9）没有额外的氨基酸残基，例如 His 标签和凝血酶切割位点。重新折叠的毒素可以抑制 rNa1.4、hNa1.5 和 hNa1.7 钠通道的失活。进一步在这些通道上进行了剂量反应实验。计算出的 EC 值分别为 rNa1.4 的 131.7±6.6nM、hNa1.5 的 454.2±50.1nM 和 hNa1.7 的 30.9±10.3μM。通道激活实验表明，rBmKM9 毒素可以将 rNa1.4 和 hNa1.5 通道的激活曲线向更负的方向移动，并呈现出典型的β-毒素特征。然而，rBmKM9 毒素对钠通道的激活特性并不相同，它会导致 rNa1.4 和 hNa1.5 通道失活的能力发生不同的变化。稳态失活的 V 值对于 rNa1.4 变得更正，对于 hNa1.5 变得更负。此外，与 rNa1.4 通道相比，rBmKM9 暴露后 hNa1.5 通道的失活恢复明显延迟。总之，这些发现强调了 rBmKM9 毒素具有 α-和 β-毒素的药理学特性，这将增加对蝎毒素经典分类的挑战。此外，钠通道的表达方法和功能信息将促进毒素的潜在应用，并有助于进一步的通道结构和功能研究。
