Meissner Gabriel Otto, de Resende Lara Pedro Túlio, Scott Luis Paulo Barbour, Braz Antônio Sérgio Kimus, Chaves-Moreira Daniele, Matsubara Fernando Hitomi, Soares Eduardo Mendonça, Trevisan-Silva Dilza, Gremski Luiza Helena, Veiga Silvio Sanches, Chaim Olga Meiri
Department of Cell Biology, Federal University of Paraná, Jardim das Américas, 81531-990, Curitiba, Paraná, Brazil.
Laboratory of Computational Biology and Bioinformatics, Federal University of ABC, Santo André, São Paulo, Brazil.
J Mol Model. 2016 Sep;22(9):196. doi: 10.1007/s00894-016-3067-0. Epub 2016 Aug 3.
Inhibitor cystine knots (ICKs) are a family of structural peptides with a large number of cysteine residues that form intramolecular disulfide bonds, resulting in a knot. These peptides are involved in a variety of biological functions including predation and defense, and are found in various species, such as spiders, scorpions, sea anemones, and plants. The Loxosceles intermedia venom gland transcriptome identified five groups of ICK peptides that represent more than 50 % of toxin-coding transcripts. Here, we describe the molecular cloning of U2-Sicaritoxin-Lit2 (U2-SCRTX-Lit2), bioinformatic characterization, structure prediction, and molecular dynamic analysis. The sequence of U2-SCRTX-Lit2 obtained from the transcriptome is similar to that of μ-Hexatoxin-Mg2, a peptide that inhibits the insect Nav channel. Bioinformatic analysis of sequences classified as ICK family members also showed a conservation of cysteine residues among ICKs from different spiders, with the three dimensional molecular model of U2-SCRTX-Lit2 similar in structure to the hexatoxin from μ-hexatoxin-Mg2a. Molecular docking experiments showed the interaction of U2-SCRTX-Lit2 to its predictable target-the Spodoptera litura voltage-gated sodium channel (SlNaVSC). After 200 ns of molecular dynamic simulation, the final structure of the complex showed stability in agreement with the experimental data. The above analysis corroborates the existence of a peptide toxin with insecticidal activity from a novel ICK family in L. intermedia venom and demonstrates that this peptide targets Nav channels.
抑制性胱氨酸结（ICKs）是一类结构肽家族，含有大量形成分子内二硫键的半胱氨酸残基，从而形成一种结状结构。这些肽参与多种生物学功能，包括捕食和防御，存在于各种物种中，如蜘蛛、蝎子、海葵和植物。中间型毒蛛（Loxosceles intermedia）毒腺转录组鉴定出五组ICK肽，占毒素编码转录本的50%以上。在此，我们描述了U2-希卡里毒素-Lit2（U2-SCRTX-Lit2）的分子克隆、生物信息学特征、结构预测和分子动力学分析。从转录组获得的U2-SCRTX-Lit2序列与μ-六聚毒素-Mg2相似，μ-六聚毒素-Mg2是一种抑制昆虫Nav通道的肽。对归类为ICK家族成员的序列进行生物信息学分析还表明，不同蜘蛛的ICKs之间半胱氨酸残基具有保守性，U2-SCRTX-Lit2的三维分子模型在结构上与μ-六聚毒素-Mg2a的六聚毒素相似。分子对接实验表明U2-SCRTX-Lit2与其可预测靶点——斜纹夜蛾电压门控钠通道（SlNaVSC）之间存在相互作用。经过200纳秒的分子动力学模拟，复合物的最终结构显示出稳定性，与实验数据一致。上述分析证实了中间型毒蛛毒液中一个新的ICK家族存在具有杀虫活性的肽毒素，并证明该肽靶向Nav通道。
