Hainantoxin-Ⅱ的分离纯化及其结构与功能分析(英文)
Isolation and characterization of Hainantoxin-II, a new neurotoxic peptide from the Chinese bird spider (Haplopelma hainanum)
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摘要: 从分布于我国海南省的海南捕鸟蛛 (Haplopelma hainanum) 毒素中, 利用阳离子交换色谱和反相高效液相色谱分离得到一种多肽神经毒素, 命名为Hainantoxin-Ⅱ(HnTx-Ⅱ)。MALDI-TOF质谱分析表明该多肽毒素相对分子质量为4 253.135, 其氨基酸序列经Edman降解测序为LFECS VSCEI EKEGN KDCKK KKCKG GWKCK FNMCV KV, 其中的6个Cys形成3对二硫键。同源性搜索表明, HnTx-Ⅱ与Huwentoxin-Ⅱ(HwTx-Ⅱ)的同源性高达91%, 仅有3个氨基酸残基不同。HwTx-Ⅱ为从虎纹捕鸟蛛中提取的杀虫肽, 该多肽具有独特的结构模体。活性分析表明HnTx-Ⅱ与HwTx-Ⅱ具有相似的生物学活性。经腹腔注射HnTx-Ⅱ, 美洲蜚蠊可被麻痹, 其ED50为16 μg/g; 而当剂量增加到60 μg/g时, 可立即杀死美洲蜚蠊, 其杀死昆虫活性明显强于HwTx-Ⅱ。此外, HnTx-Ⅱ经脑室注射可杀死小鼠, 其LD50为1.41 μg/g, 该活性却明显低于HwTx-Ⅱ。这两种多肽毒素的结构与功能的差异为进一步阐明多肽毒素的结构与功能之间的关系提供良好的研究模型。Abstract: Hainantoxin-II (HnTx-II), a novel neurotoxin, was isolated from the venom of the Chinese bird spider (Haplopelma hainanum) by cation exchange chromatography and reverse-phase HPLC. The toxin was a single chain polypeptide with calculated molecular weight of 4 253.135 obtained by mass spectrometry. The complete amino acid sequence of HnTx-II was determined by Edman degradation and found to contain 37 residues with three disulfide bonds. Results showed HnTx-II can reversibly paralyze cockroaches for several hours after intra-abdominal injection with ED50 of 16 μg/g and kill the insects immediately at a dose of 60 μg/g. It was also shown to kill mice at a LD50 value of 1.41μg/g after intracerebroventricular injection. Hainantoxin-II shares 91% sequence homology with Huwentoxin-II (HwTx-II), an insecticidal peptide from another bird spider(Haplopelma schmidti) with a unique scaffold. While HnTx-II and HwTx-II both exhibit toxic activities in insects and mammals, HnTx-II shows higher insecticidal activity and lower lethiferous activity of mammals than HwTx-II. These results help clarify structural-functional relationships ofthe polypeptide toxin.