抗脱氧雪腐镰刀菌烯醇纳米抗体随机突变与筛选  

Random Mutation and Mutant Screening of Anti-Deoxynivalenol Nanobody

作　　者：吴红静[1,2] 曹冬梅[1] 许杨[1,3] 付金衡[3] 涂追[1] 

机构地区：[1]南昌大学食品科学与技术国家重点实验室,江西南昌330047 [2]南昌大学科学技术学院,江西南昌330029 [3]南昌大学中德联合研究院,江西南昌330047

出　　处：《食品科学》2017年第10期1-5,共5页Food Science

基　　金：国家自然科学基金青年科学基金项目(31301479);江西省教育厅科学技术研究项目(GJJ151502);南昌大学科学技术学院自然科学基金项目(2013-ZR-05)

摘　　要：采用易错聚合酶链式反应技术,对来源于天然纳米抗体文库的抗脱氧雪腐镰刀菌烯醇(deoxynivalenol,DON)纳米抗体进行随机突变,通过优化和比较突变条件,构建了突变分布均匀的突变文库。噬菌体展示技术对突变文库进行了6轮筛选,获得了8种突变序列,其检测信号为野生型2.1倍。多序列比对及三维结构建模分析表明,突变序列的突变位点集中于互补决定区与框架区连接的区域,并且部分序列出现了可以形成二硫键的氨基酸残基。研究结果为抗DON纳米抗体进一步的深度突变和改造提供了线索,同时可为其他抗小分子纳米抗体的改造提供借鉴。In this work, an anti-deoxynivalenol nanobody isolated from a naive phage display library was adopted to investigate the relationship between bioactivity and the composition of amino acid residues using error prone polymerase chain reaction （epPCR） and phage display. The epPCR was firstly optimized to produce a phage display library with balanced distribution of mutation. After six rounds of panning of the mutant library, 30 out of 96 picked clones exhibited 8 different mutations, whose signal was 2.1 folds higher than that of the wild-type clone. Multi-sequence alignment analysis showed that mutant sites were prone to be found in the junction region between complementarity determining regions （CDRs） and framework regions （P^Rs）. Three-dimensional structure modeling results indicated that some mutants were capable of forming an intramolecular disulphide bond, which benefits to stabiKze the molecular structure. These data can provide a clue for further mutation and modification of anti-deoxynivalenol nanobody and also provide a guideline for the design of nanobody recognizing other low molecular weight compounds.

关 键 词：纳米抗体 脱氧雪腐镰刀菌烯醇 易错聚合酶链式反应 噬菌体展示 

分 类 号：TS201.6[轻工技术与工程—食品科学]