软骨藻酸作用蛋白质的亲和毛细管电泳筛选

doi:10.3724/SP.J.1123.2015.03026

色谱 ›› 2015, Vol. 33 ›› Issue (7): 673-677.DOI: 10.3724/SP.J.1123.2015.03026

软骨藻酸作用蛋白质的亲和毛细管电泳筛选

王晓倩¹, 高铁¹, 洪专², 屈锋¹

1. 北京理工大学生命学院, 北京 100081;
2. 国家海洋局海洋生物资源综合利用工程技术研究中心, 福建厦门 361000

收稿日期:2015-03-19 出版日期:2015-07-08 发布日期:2015-07-27
通讯作者: 屈锋
基金资助:
国家自然科学基金项目(21175011,21375008);国家"973"计划项目(2012CB910603);国家海洋局海洋生物资源综合利用工程技术研究中心开放基金(MBRCU201304).

Affinity capillary electrophoresis for screening proteins interacting with domoic acid

WANG Xiaoqian¹, GAO Tie¹, HONG Zhuan², QU Feng¹

1. School of Life Science, Beijing Institute of Technology, Beijing 100081, China;
2. Engineering Research Center of Marine Biological Resource Comprehensive Utilization, State Oceanic Administration, Xiamen 361000, China

Received:2015-03-19 Online:2015-07-08 Published:2015-07-27

摘要/Abstract

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 Δ M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。

关键词: 蛋白质, 亲和毛细管电泳, 软骨藻酸, 相互作用

Abstract:

Domoic acid (DA) is a biological neurotoxin that causes amnesic shellfish poisoning. 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 interactions between DA and nine important proteins in plasma, intestine and mitochondria were qualitatively compared by affinity capillary electrophoresis. Proteins were used as affinity ligands 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 protein 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≈trypsin≈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.

Key words: affinity capillary electrophoresis, domoic acid, interaction, protein

中图分类号:

O658

王晓倩, 高铁, 洪专, 屈锋. 软骨藻酸作用蛋白质的亲和毛细管电泳筛选[J]. 色谱, 2015, 33(7): 673-677.

WANG Xiaoqian, GAO Tie, HONG Zhuan, QU Feng. Affinity capillary electrophoresis for screening proteins interacting with domoic acid[J]. Chinese Journal of Chromatography, 2015, 33(7): 673-677.

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软骨藻酸作用蛋白质的亲和毛细管电泳筛选

Affinity capillary electrophoresis for screening proteins interacting with domoic acid

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