摘要
目的克隆表达幽门螺杆菌基因hp0410,构建12肽噬菌体展示库,对hp0410的抗原表位进行筛选和分析,为构建多表位嵌合疫苗提供理论依据。方法从幽门螺杆菌NCTC11639基因组DNA中扩增出hp0410,并克隆入pGEX-4T-1中表达。利用纯化重组蛋白筛选出特异性单抗E018,利用该单抗和M13噬菌体12肽随机肽库,构建HP0410抗原表位的抗原展示库。竞争-抑制实验验证获得的阳性噬菌体并测序,生物信息学分析HP0410的抗原表位。结果成功构建可高水平分泌型表达HP0410的原核表达系统。对13株阳性噬菌体测序后获得8个表位,其中候选表位1个。HP0410可有效抑制展示该表位的噬菌体与单抗E018结合。分析表明,获得的8个表位与HP0410同源性不高,但处于生物信息学预测的区域。结论获得8个HP0410高抗原性区域的模拟表位,其中1个为模拟单抗E018特异性结合的抗原决定基的候选表位。
Objective To study the feature of epitopes of HP0410 - a surface protein of Helicobacter pylori by using phage display technology. Methods We first constructed an expression system of HP0410 in E. coli BL21 and then we used the purified recombinant protein to scan monoclonal antibodies of Hp and got one specific monoclonal antibody - E018. The antibody was used to construct a 12 - peptides M13 phage library of HP0410. The DNA of positive phage was sequenced and the candidate was tested by competitive - inhibition array. The epitopes were analyzed by bioinformatics methods. Results We obtained 8 epitopes and a candidate epitope from the phage library by sequencing 13 phages DNA and analysis of bioinformafics. The competitive- inhibition array suggested that HP0410 could inhibit the binding between candidate phage and E018. Conclusion According to the bioinformaties analysis the epitopes may be the simulation of spatial epitopes of the high anti- genicity segments and the candidate epitope is probably the simulation of antigen determinant specifically binding to E018.
出处
《中国公共卫生》
CAS
CSCD
北大核心
2008年第9期1090-1093,共4页
Chinese Journal of Public Health
基金
广州市科技攻关项目(2007Z3-E0391)