大肠杆菌免疫蛋白OmpA生物信息学分析及表位多肽疫苗设计
|
摘要
为进一步提高大肠杆菌外膜蛋白(OmpA)的免疫效果,对大肠杆菌的OmpA进行生物信息学分析并设计其表位多肽重组疫苗。运用DNAMAN 8.0和MEGA 5.0分别对OmpA进行同源性及系统发生分析,通过ExPASy-ProtParam对OmpA的理化性质进行分析,分别采用Signal P 4.1与TMHMM Serverv.2.0对OmpA进行信号肽和跨膜结构预测,应用SOPMA和SWISS-MODEL对OmpA进行二级结构和三级结构预测,通过String进行蛋白质互作网络分析,使用BepiPred 1.0和ABCpred方法预测OmpA的B细胞抗原表位,利用神经网络法与MHC-Ⅱ类分子结合肽在线预测程序预测OmpA的CTL和Th细胞抗原表位,采用DNASTAR.Lasergene.v 7.1拼接获得优势抗原表位。进化关系结果显示,OmpA在肠道菌属间遗传进化关系较近,OmpA抗体可能为不同种肠道细菌的感染提供交叉免疫保护。OmpA为亲水性蛋白质,第1—21位氨基酸为信号肽位置,具跨膜结构。OmpA的二级结构中以无规则卷曲(45.66%)、α-螺旋(31.79%)、β-片层(16.18%)和β-转角(6.36%)为主,其三级结构为桶状结构。OmpA与par、coa蛋白之间存在互作。OmpA可能有4个B细胞抗原表位,1个CTL细胞抗原表位,1个Th细胞抗原表位,且重组获得抗原性较好的OmpA表位多肽。
In order to further improve the immune effect of Escherichia coli outer membrane protein(OmpA),and the OmpA was analyzed by bioinformatics and its epitope polypeptide recombinant vaccine was designed.The homology and phylogenetic analyses of OmpA were carried out by DNAMAN 8.0 and MEGA 5.0 softwares respectively.The physicochemical properties of OmpA were analyzed by ExPASy-ProtParam software.Signal P 4.1 and TMHMM Serverv.2.0 softwares were used to predict the signal peptide and transmembrane structures of OmpA.SOPMA and SWISS-MODEL were used to predict secondary and tertiary structures of OmpA.The protein interaction network was analyzed by String software.BepiPred 1.0 and ABCpred methods were used to predict the B cell epitopes of OmpA.Neural network method and online MHC-Ⅱ peptide binding server were used to predict the CTL and Th cell epitopes of OmpA,respectively.DNASTAR.Lasergene.v 7.1 software was used to obtain the dominant epitopes.The results of evolutionary relationship showed that OmpA had close relationship in the genetic evolution of intestinal bacteria,and OmpA might provide cross immune protection against the infection of different intestinal bacteria.OmpA was hydrophilic protein,and 1 to 21 amino acids of OmpA were the signal peptide position,which had transmembrane structure.The random coil,alpha helix,extended strand and beta-turn of OmpA secondary structure were 45.66%,31.79%,16.18% and 6.36%,respectively.The 3 D structure prediction found that OmpA was barrel structure.OmpA interacted with par protein and coa protein.The numbers of B cell epitope,CTL cell epitope and Th cell epitope of OmpA were 4,1 and 1,respectively.And the OmpA epitope polypeptides with better antigenicity was obtained by reorganization and splicing.
In order to further improve the immune effect of Escherichia coli outer membrane protein(OmpA),and the OmpA was analyzed by bioinformatics and its epitope polypeptide recombinant vaccine was designed.The homology and phylogenetic analyses of OmpA were carried out by DNAMAN 8.0 and MEGA 5.0 softwares respectively.The physicochemical properties of OmpA were analyzed by ExPASy-ProtParam software.Signal P 4.1 and TMHMM Serverv.2.0 softwares were used to predict the signal peptide and transmembrane structures of OmpA.SOPMA and SWISS-MODEL were used to predict secondary and tertiary structures of OmpA.The protein interaction network was analyzed by String software.BepiPred 1.0 and ABCpred methods were used to predict the B cell epitopes of OmpA.Neural network method and online MHC-Ⅱ peptide binding server were used to predict the CTL and Th cell epitopes of OmpA,respectively.DNASTAR.Lasergene.v 7.1 software was used to obtain the dominant epitopes.The results of evolutionary relationship showed that OmpA had close relationship in the genetic evolution of intestinal bacteria,and OmpA might provide cross immune protection against the infection of different intestinal bacteria.OmpA was hydrophilic protein,and 1 to 21 amino acids of OmpA were the signal peptide position,which had transmembrane structure.The random coil,alpha helix,extended strand and beta-turn of OmpA secondary structure were 45.66%,31.79%,16.18% and 6.36%,respectively.The 3 D structure prediction found that OmpA was barrel structure.OmpA interacted with par protein and coa protein.The numbers of B cell epitope,CTL cell epitope and Th cell epitope of OmpA were 4,1 and 1,respectively.And the OmpA epitope polypeptides with better antigenicity was obtained by reorganization and splicing.
引文
[1] GU H,LIAO Y,ZHANG J,et al.Rational design and evaluation of an artificial Escherichia coli K1 protein vaccine candidate based on the structure of OmpA[J].Frontiers in Cellular and Infection Microbiology,2018,8:172-184.
[2] HSIEH W S,YANG Y Y,LIN P H,et al.Recombinant OmpA protein fragments mediate interleukin-17 regulation to prevent Escherichia coli meningitis[J].Journal of Microbiology Immunology and Infection,2016,49(6):843-850.
[3] RAINARD P,RéPéRANTFERTER M,GITTON C,et al.Cellular and humoral immune response to recombinant Escherichia coli OmpA in cows[J].PLoS One,2017,12(10):e0187369.
[4] 卫昱君,王紫婷,徐瑗聪,等.致病性大肠杆菌现状分析及检测技术研究进展[J].生物技术通报,2016,32(11):80-92.
[5] SHARMA M,DIXIT A.Identification and immunogenic potential of B cell epitopes of outer membrane protein OmpF of Aeromonas hydrophila in translational fusion with a carrier protein[J].Applied Microbiology and Biotechnology,2015,99(15):6277-6291.
[6] GUAN Q,WANG X,WANG X,et al.Recombinant outer membrane protein A induces a protective immune response against Escherichia coli,infection in mice[J].Applied Microbiology and Biotechnology,2015,99(13):5451-5460.
[7] CONFER A W,AYALEW S.Identification of minimum carbohydrate moiety in N-glycosylation sites of brain endothelial cell glycoprotein 96 for interaction with Escherichia coli K1 outer membrane protein A[J].Microbes and infection,2014,16(7):540-552.
[8] CONFER A W,AYALEW S.The OmpA family of proteins:Roles in bacterial pathogenesis and immunity[J].Veterinary Microbiology,2013,163(3/4):207-222.
[9] NOVINROOZ A,ZAHRAEI S T,FIROUZI R,et al.In-silico design,expression,and purification of novel chimeric Escherichia coli O157:H7 OmpA fused to LTB protein in Escherichia coli[J].PLoS One,2017,12(3):e0173761.
[10] VAIPHEI S T,TANG Y,MONTELIONE G T,et al.The use of the condensed single protein production system for isotope-labeled outer membrane proteins,OmpA and OmpX in E.coli[J].Molecular Biotechnology,2011,47(3):205-210.
[11] 刘祥.溶藻弧菌附着定植因子ACFA蛋白的原核表达、抗原性鉴定及生物信息学分析[J].西南农业学报,2016,29(7):1755-1760.
[12] 俱雄,党亚锋,刘祥,等.大肠杆菌外膜蛋白OmpC的生物信息学分析及表位多肽疫苗的重组预测[J].河南农业大学学报,2017,51(1):94-100.
[13] 钟登科,黄艺珠,魏建超,等.流行性乙型脑炎病毒E蛋白多表位肽在毕赤酵母中分泌表达及其免疫原性研究[J].中国预防兽医学报,2017,39(4):296-300.
[14] 龙伟,李欣,高金燕,等.生物信息学技术在食物过敏原表位预测中的应用[J].食品科学,2014,35(3):259-263.
[15] 刘娜,孟祥英,李恒,等.结核分枝杆菌α晶体蛋白(Rv2031c)的T、B细胞抗原表位在线分析[J].细胞与分子免疫学杂志,2018,34(5):408-413.
[16] 刘祥,陈琛,陈春琳,等.猪重要感染病毒蛋白的二级结构、抗原表位分析及三联表位多肽疫苗的重组预测[J].生物学杂志,2017,34(3):18-23.
[17] 刘新生,王永录.口蹄疫病毒株AF/72结构蛋白VP1克隆及其T细胞表位预测[J].华北农学报,2010,25(5):28-32.
[18] 于娟,李红,饶华祥,等.H9N2禽流感病毒血凝素蛋白的主要特性及B/T细胞抗原表位预测分析[J].中国病原生物学杂志,2018,13(1):11-15,21.
[19] TOBIAS J,JASINSKA J,BAIER K,et al.Enhanced and long term immunogenicity of a Her-2/neu multi-epitope vaccine conjugated to the carrier CRM197 in conjunction with the adjuvant Montanide[J].Bmc Cancer,2017,17(1):118-130.
[20] 李世芳,邵军军,常惠芸.口蹄疫表位疫苗的研究进展[J].中国畜牧兽医,2017,44(2):396-401.
[21] 刘昆梅,秦玉红,奚涛,等.幽门螺旋杆菌尿素酶多表位疫苗CTB-UE的生物信息学分析及其表达[J].中国生物制品学杂志,2015,28(12):1238-1244.
[22] 杨利敏,李晶,高福,等.埃博拉病毒疫苗研究进展[J].生物工程学报,2015,31(1):1-23.
[23] 杨超,张芝晴,沈鸿霖,等.人类免疫缺陷病毒1型gp140包膜蛋白三聚体在哺乳动物细胞中的高效表达及其性质鉴定[J].病毒学报,2017,33(4):541-549.
[2] HSIEH W S,YANG Y Y,LIN P H,et al.Recombinant OmpA protein fragments mediate interleukin-17 regulation to prevent Escherichia coli meningitis[J].Journal of Microbiology Immunology and Infection,2016,49(6):843-850.
[3] RAINARD P,RéPéRANTFERTER M,GITTON C,et al.Cellular and humoral immune response to recombinant Escherichia coli OmpA in cows[J].PLoS One,2017,12(10):e0187369.
[4] 卫昱君,王紫婷,徐瑗聪,等.致病性大肠杆菌现状分析及检测技术研究进展[J].生物技术通报,2016,32(11):80-92.
[5] SHARMA M,DIXIT A.Identification and immunogenic potential of B cell epitopes of outer membrane protein OmpF of Aeromonas hydrophila in translational fusion with a carrier protein[J].Applied Microbiology and Biotechnology,2015,99(15):6277-6291.
[6] GUAN Q,WANG X,WANG X,et al.Recombinant outer membrane protein A induces a protective immune response against Escherichia coli,infection in mice[J].Applied Microbiology and Biotechnology,2015,99(13):5451-5460.
[7] CONFER A W,AYALEW S.Identification of minimum carbohydrate moiety in N-glycosylation sites of brain endothelial cell glycoprotein 96 for interaction with Escherichia coli K1 outer membrane protein A[J].Microbes and infection,2014,16(7):540-552.
[8] CONFER A W,AYALEW S.The OmpA family of proteins:Roles in bacterial pathogenesis and immunity[J].Veterinary Microbiology,2013,163(3/4):207-222.
[9] NOVINROOZ A,ZAHRAEI S T,FIROUZI R,et al.In-silico design,expression,and purification of novel chimeric Escherichia coli O157:H7 OmpA fused to LTB protein in Escherichia coli[J].PLoS One,2017,12(3):e0173761.
[10] VAIPHEI S T,TANG Y,MONTELIONE G T,et al.The use of the condensed single protein production system for isotope-labeled outer membrane proteins,OmpA and OmpX in E.coli[J].Molecular Biotechnology,2011,47(3):205-210.
[11] 刘祥.溶藻弧菌附着定植因子ACFA蛋白的原核表达、抗原性鉴定及生物信息学分析[J].西南农业学报,2016,29(7):1755-1760.
[12] 俱雄,党亚锋,刘祥,等.大肠杆菌外膜蛋白OmpC的生物信息学分析及表位多肽疫苗的重组预测[J].河南农业大学学报,2017,51(1):94-100.
[13] 钟登科,黄艺珠,魏建超,等.流行性乙型脑炎病毒E蛋白多表位肽在毕赤酵母中分泌表达及其免疫原性研究[J].中国预防兽医学报,2017,39(4):296-300.
[14] 龙伟,李欣,高金燕,等.生物信息学技术在食物过敏原表位预测中的应用[J].食品科学,2014,35(3):259-263.
[15] 刘娜,孟祥英,李恒,等.结核分枝杆菌α晶体蛋白(Rv2031c)的T、B细胞抗原表位在线分析[J].细胞与分子免疫学杂志,2018,34(5):408-413.
[16] 刘祥,陈琛,陈春琳,等.猪重要感染病毒蛋白的二级结构、抗原表位分析及三联表位多肽疫苗的重组预测[J].生物学杂志,2017,34(3):18-23.
[17] 刘新生,王永录.口蹄疫病毒株AF/72结构蛋白VP1克隆及其T细胞表位预测[J].华北农学报,2010,25(5):28-32.
[18] 于娟,李红,饶华祥,等.H9N2禽流感病毒血凝素蛋白的主要特性及B/T细胞抗原表位预测分析[J].中国病原生物学杂志,2018,13(1):11-15,21.
[19] TOBIAS J,JASINSKA J,BAIER K,et al.Enhanced and long term immunogenicity of a Her-2/neu multi-epitope vaccine conjugated to the carrier CRM197 in conjunction with the adjuvant Montanide[J].Bmc Cancer,2017,17(1):118-130.
[20] 李世芳,邵军军,常惠芸.口蹄疫表位疫苗的研究进展[J].中国畜牧兽医,2017,44(2):396-401.
[21] 刘昆梅,秦玉红,奚涛,等.幽门螺旋杆菌尿素酶多表位疫苗CTB-UE的生物信息学分析及其表达[J].中国生物制品学杂志,2015,28(12):1238-1244.
[22] 杨利敏,李晶,高福,等.埃博拉病毒疫苗研究进展[J].生物工程学报,2015,31(1):1-23.
[23] 杨超,张芝晴,沈鸿霖,等.人类免疫缺陷病毒1型gp140包膜蛋白三聚体在哺乳动物细胞中的高效表达及其性质鉴定[J].病毒学报,2017,33(4):541-549.