作为双功能抗体的SEA单链抗体片段：细菌表达系统的构建及其功能分析

SEA-scFv as a bifunctional antibody: construction of a bacterial expression system and its functional analysis.

作者信息

Sakurai N, Kudo T, Suzuki M, Tsumoto K, Takemura S, Kodama H, Ebara S, Teramae A, Katayose Y, Shinoda M, Kurokawa T, Hinoda Y, Imai K, Matsuno S, Kumagai I

机构信息

Tohoku University School of Medicine, Tohoku University, Sendai, Japan.

出版信息

Biochem Biophys Res Commun. 1999 Mar 5;256(1):223-30. doi: 10.1006/bbrc.1999.0263.

DOI:10.1006/bbrc.1999.0263

PMID:10066451

Abstract

A SEA-antibody single chain Fv (SEA-scFv) fusion protein was produced by bacterial expression system in this study. SEA-scFv has both staphylococcal enterotoxin A (SEA) effects and antibody activity directed at the epithelial mucin core protein MUC1, a cancer associated antigen. It was expressed mostly in the cytoplasm as an insoluble form. The gene product was solubilized by guanidine hydrochloride, refolded by conventional dilution method, and purified using metal-chelating chromatography. The resulting SEA-scFv fusion protein preparation was found to react with MUC1 and MHC class II antigens and had the ability to enhance cytotoxicity of lymphokine activated killer cells with a T cell phenotype against a human bile duct carcinoma cell line, TFK-1, expressing MUC1. This genetically engineered SEA-scFv fusion protein promises to be an important reagent for cancer immunotherapy.

摘要

本研究利用细菌表达系统制备了一种SEA抗体单链Fv（SEA-scFv）融合蛋白。SEA-scFv兼具葡萄球菌肠毒素A（SEA）的效应以及针对上皮粘蛋白核心蛋白MUC1（一种癌症相关抗原）的抗体活性。它主要以不溶性形式表达于细胞质中。基因产物用盐酸胍溶解，通过常规稀释法复性，并采用金属螯合层析进行纯化。结果发现，所得的SEA-scFv融合蛋白制剂可与MUC1和MHC II类抗原发生反应，并且能够增强具有T细胞表型的淋巴因子激活杀伤细胞对表达MUC1的人胆管癌细胞系TFK-1的细胞毒性。这种基因工程SEA-scFv融合蛋白有望成为癌症免疫治疗的一种重要试剂。