肉毒毒素蛋白受体sytIIN端片段基因的合成及其在大肠杆菌中的融合表达  

Synthesis of DNA Encoding Botulinum Neurotoxin Receptor syt II and Its Fusion Expression in E.coli

作　　者：史晶[1] 王慧[1] 包士中[1] 侯晓军[1] 荫俊[1] 

机构地区：[1]军事医学科学院微生物流行病研究所病原微生物生物安全国家重点实验室,北京100071

出　　处：《中国生物工程杂志》2007年第7期17-20,共4页China Biotechnology

摘　　要：旨在构建肉毒毒素蛋白受体sytIIN端片段的原核表达载体,并在大肠杆菌pMAL-c2x系统中表达MBP-Syt融合蛋白。根据GenBank中已报道的人sytII基因序列,截取N端氨基酸序列,依据大肠杆菌的偏爱密码子,设计引物人工合成全基因,将全长基因克隆至原核表达载体pMAL-c2x中,重组质粒转化大肠杆菌E.coli ER2566,IPTG诱导表达。表达产物经Amylose Resin亲和层析进行纯化,SDS-PAGE和免疫印记对其进行鉴定,并对该蛋白进行活性的初步分析,为进一步研究毒素与受体相互作用的机制奠定基础。In order to construct a prokaryotic expression vector of human receptor syt Ⅱ express recombinant MBP-Syt fusion protein in E. coli and to purify and identify its activity. N-fragment and to According to codon preference of E. coli, a DNA fragment encoding human syt Ⅱ N-fragment was synthesized, and then cloned into prokaryotic vector pMAL-c2x for sequencing. Then the recombinant plasmid pMAL-Syt was introduced into E. coli ER2566 by transformation for expression and the obtained engineered bacteria were induced by IPTG. The fusion protein was purified by amylose resin affinity chromatography and identified by SDS-PAGE and Western blot. The binding activity of the protein was determined by ELISA. It is concluded that MBP-Syt protein is of good binding activity.

关 键 词：肉毒毒素 受体 MBP融合蛋白 原核表达 

分 类 号：Q786[生物学—分子生物学]