金黄色葡萄球菌蛋白A的原核表达及生物亲和特性研究

山东大学学报(医学版) ›› 2009, Vol. 47 ›› Issue (10): 28-31.

金黄色葡萄球菌蛋白A的原核表达及生物亲和特性研究

李志会，岳盈盈，李鹏，宋楠楠，沈琮，赵元昊，孟红

山东省医学科学院基础医学研究所山东省现代医用药物与技术重点实验室，济南 25006

收稿日期:2008-12-18 出版日期:2009-10-16 发布日期:2009-10-16
通讯作者: 孟红(1960- )，女，研究员，硕士生导师，从事分子病毒学研究。Email：menghyky@yahoo.com.cn
作者简介:李志会(1978- )，男，助理研究员，硕士，主要从事分子病毒学研究。

Prokaryotic expression of Staphylococal Protein A and
initial study of the affinity

LI Zhihui， YUE Yingying， LI Peng， SONG Nannan， SHEN Cong， ZHAO Yuanhao， MENG Hong

Institute of Basic Medicine，Shandong Academy of Medical Sciences， Key

Received:2008-12-18 Online:2009-10-16 Published:2009-10-16

摘要/Abstract

摘要：

目的构建金黄色葡萄球菌蛋白A (SPA)基因片段的表达载体，并在大肠杆菌表达系统中进行表达，探讨该重组蛋白与不同来源IgG结合的生物学特性。方法利用PCR法扩增SPA基因片段，将该段基因克隆到pET30a(+)表达载体，转化感受态BL21大肠杆菌细胞，IPTG诱导表达,利用SDSPAGE及Westenblot鉴定目的蛋白。His亲和层析纯化后分别与人、小鼠、兔、猪及山羊血清反应，分析其与不同来源IgG的亲和关系。结果构建的pET30a(+)SPA表达载体在大肠杆菌BL21中可表达SPA蛋白，且发现其与人及小鼠源IgG具有较高亲和性，与兔及猪源IgG亲和性较低。结论该重组蛋白SPA与人及小鼠源免疫球蛋白IgG具有较高亲和性，为下一步应用于SPAELISA打下基础。

关键词: 葡萄球菌蛋白A, 原核表达, 亲和性

Abstract:

To construct an expression plasmid and express Staphylococcus aureus Protein A(SPA) in a prokaryotic expression system. The initially study concerned the affinity of purified SPA with IgGs from different sorts of animals. MethodsPCR was used to amplify the target gene with the genome of Staphylococcus aureus as a template. The recombination plasmid pET30a(+)SPA was constructed and transformed into E.coli BL21 competent cells. The recombinant protein was expressed with induction of IPTG and identified by SDSPAGE and Westernblot followed by purification with His·Bind Protein Purification Kit. After that ELISA was used to study the affinity of purified SPA with different IgGs from humans, rabbits, mice, swine and goats. ResultsThe expression plasmid pET30a(+)SPA was constructed successfully and the objective protein was expressed. The recombination protein showed high affinity with IgGs from humans and mice but low affinity with IgGs from rabbits and swine. ConclusionThe recombination protein has high affinity with IgGs from humans and mice but low affinity with IgGs from rabbits and swine, and so lays a foundation for SPAELISA.

Key words: Staphylococal Protein A； Prokaryotic expression； Affinity

中图分类号:

. 金黄色葡萄球菌蛋白A的原核表达及生物亲和特性研究[J]. 山东大学学报(医学版), 2009, 47(10): 28-31.

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