海南单柄蛛毒素杀虫肽HNTXa的分离与性质分析

Purification and analysis of an insecticidal peptide from the venom of Ornithoctonus hainana

为了从海南单柄蛛Ornithoctonus hainana毒素中获得对哺乳动物无毒而对昆虫有毒杀作用的单一蛋白组分,应用反相层析和阳离子交换对海南单柄蛛粗毒素进行分离纯化。分别通过胸腔注射、腹腔注射和饲喂的方法测定各毒素组分对蟋蟀、小鼠和棉铃虫的毒性作用,并采用改进的寇式法计算LD50。分离得到一种新的杀虫肽,命名为海南单柄蛛毒素a(Hainana toxica,HNTXa),聚丙烯酰胺等点聚焦电泳(IEF-PAGE)分析其等电点为8.15～8.45。杀虫肽HNTXa对棉铃虫和蟋蟀有较强的抑制作用。棉铃虫经饲喂浸沾HNTXa的棉花叶片72h后,死亡率超过30%;HNTXa对蟋蟀的LD50值为58μg/g,LD50的95%置信区间为55～62μg/g;HNTXa对哺乳动物小鼠无明显毒性。

In order to obtain a single protein component from Ornithoctonus hainana crude venom, which had toxicity on insects but didn＇ t have toxicity on mammals, reverse phase chromatography （RPC） and cation exchange chromatography were used to isolate the venom. Intrapleural injection, intraperitoneal injection and feeding method were used to measure toxicity of each toxin component on crickets, mouse and bollworm, respectively. Modified Karber＇ s method was used to calculate LD50. A new insecticidal peptide was separated from the venom and named hainana toxin a （HNTXa）. The map of IEF-PAGE indicates its isoelectric point is between 8.15 and 8.45. Insecticidal peptide HNTXa has strong inhibition to cotton bollworm （Helicoverpa armigera） and a cricket. Cotton bollworm was fed on cotton leaves with HNTXa and mortality rate exceed 30% 72 hours later. The LD50 values of this toxin acting on cricket is 58μg/g and the 95% confidence interval of LD50 was 55 -62μg/g. HNTXa didn＇ t have obvious toxicity on mammals.