α-溶血素的表达及其纳米孔的制备  

The Expression of α-Hemolysin and the Preparation of Its Nanopore

作　　者：卓丽霞[1] 王莹[2] 张春萍[2] 段静[2] 张亚妮[1] ZHUO Li-xia WANG Ying ZHANG Chun-ping DUAN Jing ZHANG Ya-ni(The College of Life Science, Northwest University, Xi＇an 710069, China College of Chemistry ＆ Mateirals Secience, Northwest University, Xi＇ an 710069, China)

机构地区：[1]西北大学生命科学学院,西安710069 [2]西北大学化学与材料科学学院,西安710069

出　　处：《中国生物工程杂志》2017年第1期53-57,共5页China Biotechnology

基　　金：国家自然科学基金(21175105);中国博士后基金(2012M521795)资助项目

摘　　要：α-溶血素(α-hemolysin,αHL)是金黄色葡萄球菌分泌的一种水溶性穿孔毒素,在双脂膜上可组装成结构稳定的七聚体蛋白纳米孔,是理想的单分子检测器件。以大肠杆菌BL21(DE3)p Lys S作为表达宿主菌株诱导表达野生型和单位点突变型α-溶血素,超滤膜分子截留法纯化蛋白质,使用兔血红细胞膜将αHL单体组装成七聚体蛋白纳米孔,并在人工脂双层膜上构建适于单分子分析的分子器件。通过溶血实验和单通道电流记录,证实αHL溶血活性良好,组装的αHL七聚体纳米孔在双脂膜上打孔能力和离子通透性良好,结构稳定,可作为理想的单分子检测器件。此方法为大量制备αHL单体及其在单分子分析和应用研究方面奠定了基础。α-hemolysin（ αHL） is a channel-forming toxin released by Staphylococcus aureus,which can form a transmembrane heptamers nanopore on eukaryotic cell membranes and other lipid bilayers. The αHL nanopore has been popularly used as a single molecule biosensor. In the present study,αHL was expressed in the host strain BL21（ DE3） p Lys S and purified by using molecular weight cut-off of ultrafiltration membranes. αHL heptameric nanopore was formed on rabbit red blood cell memberanes and characterized by using patch clamp technique and single channel current recording. αHL heptameric nanopore showed a stable structure and good ion permeability on lipid bilayer. It can be used as a stable single-molecule detection device. These results make a useful work for the preparation of α-hemolysin heptameric nanopore and the further exploring in the filed of single-molecule detection.

关 键 词：α-溶血素 蛋白纳米孔 单通道电流 单分子检测器件 

分 类 号：Q819[生物学—生物工程]