Expression of Chimeric Protein of Norovirus P Domain and Neutralizing Epitopes of EV71 in Prokaryotic <em>Escherichia coli</em>

doi:10.13523/j.cb.20170101

China Biotechnology

2017, Vol. 37

Issue (1): 1-6 DOI: 10.13523/j.cb.20170101

Expression of Chimeric Protein of Norovirus P Domain and Neutralizing Epitopes of EV71 in Prokaryotic Escherichia coli

LI Chao, LIU Bo, TAO Yu-fen, LI Xin-tong, LIU Jian-sheng, LIU Hong-qi

Infection and Immunity Laboratory, Institute of Medical Biology, Chinese Academy of Medical Sciences & Peking Union Medical College, Kunming 650118, China

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Abstract

Objective:To construct recombinant plasmids and express chimeric proteins of neutralizing epitopes of Human Enterovirus 71 and Norovirus P domain. Methods:Nucleotide sequences of the neutralizing epitopes of EV71 were designed and optimized according to their reported sequences of amino acids and the codon bias usage of E. coli for expression. These nucleotide fragments were cloned into the plasmid vector containing norovirus P domain and GST tag via the loop2 of P domain, which was confirmed by sequencing. The constructs were transformed into the bacteria BL21(DE3) and induced with IPTG for protein expression. The GST-fused proteins were purified via GST affinity beads. Immunoblot analysis was used to detect the GST tag and evaluate the immunogenicity of chimeric proteins. Results:Seven recombinant plasmids were constructed successfully as confirmed by sequencing. The expressed 7 chimeric proteins were expressed and soluble in E. coli, which was determined by SDS-PAGE and Commassie blue staining. Immunoblot analysis via the anti-GST antibody revealed the expression of GST-fused proteins. Further analysis of immunogenicity showed that all 7 chimeric proteins could react with anti-NoV P domain antibody. Five out of 7 chimeric proteins could recognize anti-EV71 polyclonal antibody except two epitopes of SP55 and SP28 chimeric proteins. Conclusion:The chimeric proteins of NoV P domain and EV71 neutralizing epitopes were successfully expressed and antigenic, which lays a solid foundation for developing the bivalent subunit vaccine and the detection kit for NoV and EV71 viruses.

Key words： Enterovirus 71 Norovirus Neutralizing epitopes Subunit vaccine

Received: 04 August 2016 Published: 25 January 2017

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LI Chao, LIU Bo, TAO Yu-fen, LI Xin-tong, LIU Jian-sheng, LIU Hong-qi. Expression of Chimeric Protein of Norovirus P Domain and Neutralizing Epitopes of EV71 in Prokaryotic Escherichia coli. China Biotechnology, 2017, 37(1): 1-6.

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https://manu60.magtech.com.cn/biotech/10.13523/j.cb.20170101 OR https://manu60.magtech.com.cn/biotech/Y2017/V37/I1/1

[1] Kapikian A Z, Wyatt R G, Dolin R, et al. Visualization by immune electron microscopy of a 27-nm particle associated with acute infections nonbacterial gastroenteritis. Journal of Virology, 1972, 10(5):1075-1081.
[2] Dolin R. Noroviruses-challenges to control. N Engl J Med, 2007, 357(11):1072-1073.
[3] Yang F, Ren L L, Xiong Z H, et al. Enterovirus 71 outbreak in the People's Republic of China in 2008. Journal of Clinical Microbiology, 2009, 47(7):2351-2352.
[4] Chen S C, Chang H L, Yan T R, et al. An eight-year study of epidemiologic features of Enterovirus 71 infection in Taiwan. Am J Trop Med Hyg, 2007, 77(1):188-191.
[5] Prasad B V V, Hardy M E, Dokland T, et al. X-ray crystallographic structure of the Norwalk virus capsid. Science, 1999, 286(5438):287-290.
[6] Cao S, Lou Z Y, Tan M, et al. Structural basis for the recognition of blood group trisaccharides by norovirus. Journal of Virology, 2007, 81(11):5949-5957.
[7] Tan M, Fang P A, Chachiyo T, et al. Noroviral P particle:structure, function and applications in virus-host interaction. Virology, 2008, 382(1):115-123.
[8] Tan M, Jiang X. The P domain of norovirus capsid protein forms a subviral particle that binds to histo-blood group antigen receptors. Journal of Virology, 2005, 79(22):14017-14030.
[9] Fang H, Tan M, Xia M, et al. Norovirus P particle efficiently elicits innate, humoral and cellular immunity. PLoS One, 2013, 8(4):e63269.
[10] Tan M, Huang P W, Xia M, et al. Norovirus P particle, a novel platform for vaccine development and antibody production. Journal of Virology, 2011, 85(2):753-764.
[11] Liu C C, Chou A H, Lien S P, et al. Identification and characterization of a cross-neutralization epitope of Enterovirus 71. Vaccine, 2011, 29(26):4362-4372.
[12] Foo D G W, Alonso S, Phoon M C, et al. Identification of neutralizing linear epitopes from the VP1 capsid protein of Enterovirus 71 using synthetic peptides. Virus Research, 2007, 125(1):61-68.
[13] Li Y X, Zhao H, Cao R Y, et al. Recombinant tandem multi-linear neutralizing epitopes of human Enterovirus 71 elicited protective immunity in mice. Virology Journal, 2014, 11(1):79-86.
[14] 高文超, 国泰, 王玉琳. EV71抗原表位的研究方法及研究进展. 中国生物制品学杂志, 2014, 27(6):852-855. Gao W C, Guo T, Wang Y L. Method for and advance in research on neutralizing epitope of EV71. Chin J Biologicals, 2014, 27(6):852-855.
[15] Tan M, Jiang X. Norovirus P particle, a subviral nanoparticle for vaccine development against norovirus, rotavirus and influenza virus. Nanomedicine, 2012, 7(6):889-897.
[16] Xia M, Tan M, Wei C, et al. A candidate dual vaccine against influenza and noroviruses. Vaccine, 2011, 29(44):7670-7677.
[17] Wang L Y, Cao D J, Wei C, et al. A dual vaccine candidate against norovirus and hepatitis E virus. Vaccine, 2014, 32(4):445-452.
[18] Li P, Zheng Q S, Wang Q, et al. Immune responses of recombinant adenoviruses expressing immunodominant epitopes against Japanese encephalitis virus. Vaccine, 2008, 26(46):5802-5807.
[19] Su C X, Duan X G, Wang X Q, et al. Heterologous expression of FMDV immunodominant epitopes and HSP70 in P. pastoris and the subsequent immune response in mice. Veterinary Microbiology, 2007, 124(3-4):256-263.

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