从噬菌体表面展示环状肽肽库中筛选葡萄球菌B型肠毒素抑制剂.docxVIP

从噬菌体表面展示环状7肽肽库中筛选葡萄球菌B型肠毒素抑制剂 作者：李韩平，周宏，郑玉玲，邓小红，马茹，姜永强 【关键词】 葡萄球菌B型肠毒素 Screening inhibitor of Staphylococcal enterotoxin B from random circular 7mer peptide library displayed on phage 【Abstract】 AIM: To obtain the specific peptide which can bind with SEB and inhibit SEBs enterotoxin activity by panning from the random circular 7mer polypeptide libraries. METHODS: The affinity of peptide and the competitive activity of the synthetic peptide with that of antiSEB monoclonal antibody were detected respectively by PhageELISA and competition ELISA. Animal experiment was performed to assess the suppressive effect of the screened peptide on the toxicity of SEB and its enterotoxin. RESULTS: The circular 7mer peptide obtained by panning suppressed the multiplication of spleen lymphocytes in mice. At the ratio of 160:1 , the peptide protected the little cats from attack of enterotoxin and the survival rate of mice treated with the screened peptide was significantly higher compared with that of the control group. CONCLUSION: The obtained peptide specifically combines with SEB and suppresses its enterotoxin activity, which paves the way for the development of SEB inhibitors. 【Keywords】 staphylococcal enterotoxin B; peptide library; screening; inhibitor 【摘要】 目的： 拟通过生物淘选从噬菌体表面展示环状7肽肽库中筛选出与葡萄球菌B型肠毒素结合并能抑制该毒素毒性效应的特异性短肽. 方法： 采用PhageELISA来鉴定所得目的肽的亲和性；利用竞争ELISA研究合成肽与SEB mAb竞争结合SEB的情况；通过动物实验考察其抑制SEB的超抗原特性和肠毒活性情况. 结果： 筛选所得短肽在一定浓度范围内可以抑制SEB对鼠脾淋巴细胞的激活；合成肽与SEB质量比160∶1下，合成肽可较好地抑制SEB对乳猫的肠毒活性，并对SEB引起的小鼠致死具有明显保护作用. 结论： 得到了能与SEB特异结合并能抑制SEB超抗原特性和肠毒活性的短肽. 【关键词】 葡萄球菌B型肠毒素；肽库;筛选;抑制剂 　0引言 　　 　　葡萄球菌B型肠毒素［1］是超抗原家族的主要成员之一，该毒素可以造成食物中毒，严重时甚至导致致死性休克. SEB中毒后缺乏特异的治疗手段，因此研究SEB的特异高效抑制剂具有重大的意义. 我们利用固相筛选方法，即以SEB作为亲和靶标蛋白，从环状7mer肽噬菌体文库中筛选到与SEB结合，并能抑制该毒素毒性效应的噬菌体克隆. 结果表明，所筛选环状7mer肽在细胞水平上可抑制SEB对小鼠脾细胞的激活，在整体动物水平上对SEB导致的毒性具有一定保护作用. 1材料和方法 材料 环状7mer肽噬菌体文库为New England BioLabsInc产品; M13噬菌体单链DNA提取试剂盒为Qiagen公司产品； SEB单克隆抗体由本室保存; DGa