抗A型肉毒毒素人源三结构域抗体的重组设计与表达  

Recombinant design and expression of human three-domain antibody against BoNTa

作　　者：王慧[1] 荫俊[1] 史晶[1] 孟露露[1] 李佩真[1] 

机构地区：[1]军事医学科学院微生物流行病研究所,北京100071

出　　处：《微生物学报》2005年第2期223-225,共3页Acta Microbiologica Sinica

基　　金：国家"973项目"(2 0 0 2CB5 13 2 0 5 )~~

摘　　要：以获得的抗A型肉毒毒素人源单链抗体ScFvB17为模板 ,PCR扩增抗体ScFv基因的重链可变区 (VH)和轻链可变区 (Vκ) ,以重链恒定区 1(CH1) 5′端 12个氨基酸的序列作为连接肽 ,构建成三结构域抗体分子 :VH_连接肽_Vκ(VH Vκ)。重组VH Vκ抗体分子在大肠杆菌中得到高效表达 ,其表达量占菌体总蛋白质的 34%以上。表达的蛋白质在菌内形成包含体 ,采用镍柱金属鏊合层析法对该包含体进行纯化 ,得到了纯度高达 95 %的VH Vκ。ELISA等实验结果显示 ,重组设计并制备的三结构域抗体蛋白VH Vκ不但可以特异识别毒素抗原 ,具有与原母本单链抗体ScFv相同的抗原特异性 ,而且具有了更高的相对亲和力和稳定性。VH and Vκ genes were amplified from human ScFv B17 specific against botulinum neurotoxin serotype A(BoNTa). Gene sequence encoding 5′-terminal 12 amino acid of heavy chain constant region CH1, as a linker, linked VH and Vκ to construct a new three-domain antibody molecule VH/Vκ. VH/Vκ was expressed at high level over 34 of total host cell proteins in E.coli. Recombinant protein were purified up to 95 by affinity column. As a result, recombinant VH/Vκ could recognize and bind specific to BoNTa in ELISA. However, comparing with its parent ScFv, VH/Vκ has higher relative affinity and stability.

关 键 词：A型肉毒毒素 三结构域抗体 表达 中和活性 

分 类 号：R392[医药卫生—免疫学]