融合蛋白接头linker在细粒棘球蚴重组疫苗EgG1Y162-2(4)中的选择设计  

Selective design of linker fusion protein connector in echinococcus granulosus recombinant vaccine EgG1Y162-2(4)

作　　者：郑佳 赵商岐 李艳敏 周彦霞 丁剑冰[1] 周晓涛(指导)[1] ZHENG Jia;ZHAO Shangqi;LI Yanmin;ZHOU Yanxia;DING Jianbing;ZHOU Xiaotao(Basic Medical College of Xinjiang Medical University,Urumqi 830011,China)

机构地区：[1]新疆医科大学基础医学院,乌鲁木齐830011

出　　处：《中国免疫学杂志》2023年第9期1913-1921,共9页Chinese Journal of Immunology

基　　金：国家自然科学基金项目(81760656);新疆维吾尔自治区自然科学基金项目(2018D01C157)。

摘　　要：目的:分析蛋白EgG1Y162-2通过不同长度接头序列连接后形成的肽段四聚体蛋白EgG1Y162-2(4)的氨基酸序列特征,明确其理化性质及三级结构,预测并对比经不同长度柔性接头连接后其优势抗原表位变化,为增强重组疫苗免疫原性选取最理想的linker序列。方法:利用生物信息学方法选择GSGGSG、GGGGSGGG和GSGGSGGGSGGSGGG 3种linker序列进行连接设计EgG1Y162-2(4)重组蛋白疫苗。在线软件ProtParam分析其理化性质;SOPMA在线数据库对设计有不同linker序列的重组蛋白二级结构进行预测分析;I-TASSER在线软件预测所设计重组疫苗的三级结构,并使用SYFPEITHI、IEDB等软件对其T/B细胞抗原表位进行预测。结果:经预测通过GSGGSG、GGGGSGGG、GSGGSGGGSGGSGGG 3种linker序列连接的EgG1Y162-2(4)均是稳定蛋白且具有亲水性并得到其二级结构特点。设计的重组疫苗EgG1Y162-2(4)通过生物信息学方法预测得知通过GSGGSG连接的EgG1Y162-2(4)中α螺旋占8.50%,β-转角占16.67%,无规则卷曲约占40.48%,延伸链约占34.35%,该蛋白有8个T/B联合表位,前后串联的EgG1Y162-2蛋白不仅能够正常折叠,且与EgG1Y162-2蛋白相比T/B抗原表位未发生偏移。三级结构显示通过linker序列GSGGSG串联的4个蛋白EgG1Y162-2均能正常表达,而通过GGGGSGGG、GSGGSGGGSGGSGGG串联的蛋白EgG1Y162-2(4)表位发生了一定程度右移。结论:利用GSGGSG 6个氨基酸连接EgG1Y162-2蛋白构成的EgG1Y162-2(4)的T/B联合表位高度重合,表位未发生迁移说明蛋白EgG1Y162-2(4)通过这种方式连接后几乎未对EgG1Y162-2蛋白优势表位造成影响,并通过重复蛋白表位数增强疫苗免疫应答效果,制备有效的棘球蚴病表位疫苗。Objective:To analyze amino acid sequence characteristics of peptide tetramer protein EgG1Y162-2(4)formed after protein EgG1Y162-2 was linked by different lengths of joint sequences were,to clarify its physical and chemical properties and three-level structure,to predict and compare changes of its dominant antigen epitopes after linked by flexible joints of different lengths,and to select optimal linker sequence to enhance immunogenicity of recombinant vaccine.Methods:Bioinformatics methods were used to select GSGGSG,GGGGSGGG and GSGGSGGGSGGSGGG linker sequences for linking to design EgG1Y162-2(4)recombinant protein vaccine.Physicochemical properties were analyzed by online software ProtParam.SOPMA online database was used to predict secondary structure of recombinant proteins with different linker sequences.Tertiary structure of recombinant vaccine was predicted by I-TASSER online software,and its T/B cell epitopes were predicted by SYFPEITHI,IEDB and other softwares.Results:EgG1Y162-2(4)linked by GSGGSG,GGGGSGGG and GSGGSGGGSGGSGGG linker sequences were predicted to be stable proteins with hydrophilic properties,and its secondary structure characteristics were obtained.Recombinant vaccine EgG1Y162-2(4)was designed,which predicted by bioinformatics method thatα-helix of was 8.50%,β-folding accounted for 16.67%,irregular curl accounted for 40.48%and extended chain accounted for 34.35%by GSGGSG,which had 8 T/B combined epitopes.Back-to-back series EgG1Y162-2 protein not only folded normally,but also did not shift T/B epitope compared with EgG1Y162-2 protein.Tertiary structure showed that four proteins EgG1Y162-2 connected by linker sequence GSGGSG could be expressed normally,while protein EgG1Y162-2(4)epitope connected by GGGGSGGG and GSGGSGGGSGGSGGG had a certain degree of right shift.Conclusion:T/B joint epitope of EgG1Y162-2(4),which is formed by GSGGSG six amino acids to connect EgG1Y162-2 protein,highly overlapped,and no epitope migration occurred,indicating that protein EgG1Y162-2(4)is connected in this way

关 键 词：细粒棘球蚴 EgG1Y162-2 融合蛋白接头 

分 类 号：R383.3[医药卫生—医学寄生虫学]