软骨藻酸作用蛋白质的亲和毛细管电泳筛选

摘要

基于亲和毛细管电泳，定性比较了血浆、肠道及细胞线粒体中所选的9种重要功能蛋白质与神经毒性生物毒素———软骨藻酸（ domoic acid，DA）的相互作用。以 DA淌度比变化量ΔM 与蛋白质浓度 L 作图，根据斜率值大小可比较 DA与蛋白质作用的相对强弱。结果如下：可与 DA相互作用的有6种蛋白质，作用强弱为：人凝血酶＞细胞色素 C≈胰蛋白酶≈免疫球蛋白 E（ Ig E）≈核糖核酸酶 A＞λ核酸外切酶；而铁蛋白、转铁蛋白、凝集素与 DA未表现出相互作用和亲和力。实验表明，亲和毛细管电泳具有高效、快速、所需样品量低的优点，可用于DA作用靶蛋白的筛选，为 DA的毒性机制研究和毒性防护提供基础信息。%Domoic acid( DA)is a biological neurotoxin that causes amnesic shellfish poison-ing. Study of the interactions between DA and important functional proteins contributes to understand the toxicity mechanism of DA to biological macromolecules. In this paper,the inter-actions between DA and nine important proteins in plasma,intestine and mitochondria were qualitatively compared by affinity capillary electrophoresis. Proteins were used as affinity lig-ands while DA as the affinity receptor. Proteins with the concentrations of 0,0. 2,0. 4,0. 6, 0. 8 μmol/L were added in the electrophoresis buffer and the migration times of 0. 2 mg/mL DA were detected. Then the linear graphs of the variation of DA mobility ratio(ΔM)with the pro-tein mass concentration( L)were drawn. According to the slope value,the relative strength of the interactions between DA and proteins was compared. The results showed that six proteins can interact with DA and the relative strength order was human thrombin>cytochrome C≈tryp-sin≈immunoglobulin E( Ig E)≈ribonuclease A>λ exonuclease,while ferritin,transferrin and lectin had no affinity with DA. With the advantages of high efficiency,fast analysis and less sample consumption,affinity capillary electrophoresis is a convenient method for screening DA target proteins,which will provide basic information for the toxic mechanism and defence of DA.