日本血吸虫MBLAC1基因的克隆及生物信息学分析
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摘要
目的克隆日本血吸虫金属β内酰胺酶结构域蛋白1(Metallo-beta-lactamases domain-containing protein 1,MBLAC1)基因,并研究其编码蛋白的生物学特性。方法以成虫虫体cDNA为模板,利用PCR技术扩增Sj MBLAC1基因,并通过生物信息学技术分析该基因编码蛋白的结构特征。结果 Sj MBLAC1扩增基因片段大小为711bp,编码236个氨基酸,预测蛋白质分子量约为26kD,理论等电点为4.84。该蛋白的第7~222位氨基酸为保守结构域,属于MBL超级家族;不存在跨膜结构域及信号肽;具有一个N-糖基化位点,在第186位氨基酸;二级结构中包含α-螺旋8.90%、β-折叠27.97%、无规则卷曲区域63.14%,推测Sj MBLAC1蛋白具有9个优势B细胞抗原表位。结论成功克隆了Sj MBLAC1基因并对其编码蛋白进行了生物信息学分析,从而为开展该蛋白的生物学功能研究及筛选抗血吸虫病疫苗候选分子提供了基础。
In order to clone the gene of metallo-beta-lactamases domain-containing protein 1 fromSchistosoma japonicum(Sj MBLAC1),and analyze the biological characteristics of the coding protein,the Sj MBLAC1 gene was amplified by the polymerase chain reaction(PCR)technique with adult worm cDNA as a template,and the structural features of the coding protein were analyzed by the bioinformatics technique.Results showed that the full-length of Sj MBLAC1 gene was 711 bp and it encoded a protein of 236 amino acids with a predicted molecular weight of 26 kDa and an isoelectric point of 4.84.It contained a conserved domain from the 7 th to the 222 nd amino acids which belonged to the MBL super family,and an N-glycosylation site at the 186 th amino acid without the signal peptide and the transmembrane domain.In addition,its secondary structures comprised8.90% α-helices,27.97%β-sheets,and 63.14% random coils.At last,it was speculated that the Sj MBLAC1 protein had 9 dominant B cell antigen epitopes.It is concluded that Sj MBLAC1 was successfully cloned and analyzed by the bioinformatics technique.This study will provide the foundation for proceeding further research on the biological function of Sj MBLAC1 and permit the screening new vaccine candidates against schistosomiasis.
In order to clone the gene of metallo-beta-lactamases domain-containing protein 1 fromSchistosoma japonicum(Sj MBLAC1),and analyze the biological characteristics of the coding protein,the Sj MBLAC1 gene was amplified by the polymerase chain reaction(PCR)technique with adult worm cDNA as a template,and the structural features of the coding protein were analyzed by the bioinformatics technique.Results showed that the full-length of Sj MBLAC1 gene was 711 bp and it encoded a protein of 236 amino acids with a predicted molecular weight of 26 kDa and an isoelectric point of 4.84.It contained a conserved domain from the 7 th to the 222 nd amino acids which belonged to the MBL super family,and an N-glycosylation site at the 186 th amino acid without the signal peptide and the transmembrane domain.In addition,its secondary structures comprised8.90% α-helices,27.97%β-sheets,and 63.14% random coils.At last,it was speculated that the Sj MBLAC1 protein had 9 dominant B cell antigen epitopes.It is concluded that Sj MBLAC1 was successfully cloned and analyzed by the bioinformatics technique.This study will provide the foundation for proceeding further research on the biological function of Sj MBLAC1 and permit the screening new vaccine candidates against schistosomiasis.
引文
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[19]梁瑾,王靖飞.B细胞表位预测方法研究进展[J].生命科学,2009,21(2):320-323.
[2]张利娟,徐志敏,钱颖骏,等.2015年全国血吸虫病疫情通报[J].中国血吸虫病防治杂志,2016,28(6):611-617.DOI:10.16250/j.32.1374.2016246
[3]Ismail M,Metwally A,Farghaly A,et al.Characterization of isolates of Schistosoma mansoni from Egyptian villagers that tolerate high doses of praziquantel[J].Am J Trop Med Hyg,1996,55(2):214-218.DOI:10.4269/ajtmh.1996.55.214
[4]Stelma FF,Talla I,Sow S,et al.Efficacy and side effects of praziquantel in an epidemic focus of Schistosoma mansoni[J].Am J Trop Med Hyg,1995,53(2):167-170.DOI:10.4269/ajtmh.1995.53.167
[5]Pearce EJ.Progress towards a vaccine for schistosomiasis[J].Acta Trop,2003,86(2/3):309-313.DOI:10.1016/S0001-706X(03)00062-7
[6]Jones MK,Gobert GN,Zhang L,et al.The cytoskeleton and motor proteins of human schistosomes and their roles in surface maintenance and host-parasite interactions[J].Bioessays,2004,26(7):752-765.DOI:10.1002/bies.20058
[7]van Balkom BW,van Gestel RA,Brouwers JF,et al.Mass spectrometric analysis of the Schistosoma mansoni tegumental sub-proteome[J].J Proteome Res,2005,4(3):958-966.DOI:10.1021/pr050036w
[8]Zhang M,Fu Z,Li C,et al.Screening diagnostic candidates for schistosomiasis from tegument proteins of adult Schistosoma japonicum using an immunoproteomic approach[J].PLoS Negl Trop Dis,2015,9(2):e0003454.DOI:10.1371/journal.pntd.0003454
[9]陈照强,刘一方,朱宁,等.金属β-内酰胺酶的研究进展[J].国外医药抗生素分册,2011,32(3):111-115.
[10]吕晴,单思,王丽君,等.猪附红细胞体ORF5蛋白的空间结构与B细胞抗原表位预测[J].中国人兽共患病学报,2015,31(3):251-254.DOI:10.3969/j.issn.1002-2694.2015.03.014
[11]Fonseca CT,Braz Figueiredo Carvalho G,Carvalho Alves C,et al.Schistosoma tegument proteins in vaccine and diagnosis development:an update[J].J Parasitol Res,2012,2012(3):541268.DOI:10.1155/2012/541268
[12]Han ZG,Brindley PJ,Wang SY,et al.Schistosoma genomics:new perspectives on schistosome biology and host-parasite interaction[J].Annu Rev Genomics Hum Genet,2009,10(10):211-240.DOI:10.1146/annurev-genom-082908-150036
[13]Molehin AJ,Gobert GN,McManus DP.Serine protease inhibitors of parasitic helminths[J].Parasitology,2012,139(6):681-695.DOI:10.1017/S0031182011002435
[14]Dinguirard N,Yoshino TP.Potential role of a CD36-like class B scavenger receptor in the binding of modified low-density lipoprotein(acLDL)to the tegumental surface of Schistosoma mansoni sporocysts[J].Mol Biochem Parasitol,2006,146(2):219-230.DOI:10.1016/j.molbiopara.2005.12.010
[15]Wenzel NF,Carenbauer AL,Pfiester MP,et al.The binding of iron and zinc to glyoxalase II occurs exclusively as di-metal centers and is unique within the metallo-β-lactamase family[J].J Biol Inorg Chem,2004,9(4):429.DOI:10.1007/s00775-004-0535-2
[16]Zhang M,Han Y,Zhu Z,et al.Cloning,expression,and characterization of Schistosoma japonicum tegument protein phosphodiesterase-5[J].Parasitol Res,2012,110(2):775-786.DOI:10.1007/s00436-011-2552-8
[17]王家红,童玥,朱玥,等.蛋白质糖基化的研究进展[J].药物生物技术,2011,18(1):77-80.
[18]马凡舒,张蕾,王洋,等.B细胞抗原表位预测方法的研究进展[J].中国畜牧兽医,2016,43(1):63-67.
[19]梁瑾,王靖飞.B细胞表位预测方法研究进展[J].生命科学,2009,21(2):320-323.