Preparation and identification of Norovirus GⅡ.17 virus-like particles

Abstract:

[Background] Norovirus, featuring genetic diversity, is a major foodborne initiator of acute gastroenteritis in human worldwide. GⅡ.17, the most destructive genotype, is a novel variant emerging in Asia from 2014 through 2015 and has been predominant in Asia. Virus-like particles (VLPs) of Norovirus, prepared by genetic engineering, are safe with high immunogenicity, which are promising candidate vaccine of Norovirus. [Objective] This paper aims to prepare GⅡ.17 VLPs in Escherichia coli. [Methods] The gene encoding GⅡ.17 capsid protein VP1 was synthesized and cloned into pET28a-MsyB vector. The recombinant plasmid was transformed into E. coli BL21(DE3) and expressed under the induction of IPTG. The fusion tags were removed by tobacco etch virus (TEV) protease, followed by purification with His-Tag nickel column to yield the natural VP1 protein. SDS-PAGE, Western blot, and transmission electron microscope were employed to determine the reactogenicity and structure of VP1 protein. [Results] MsyB-VP1 can be expressed in soluble form, and the optimum conditions are as follows:IPTG at 0.8 mmol/L, and 37℃ for 8 h. The purified VP1 protein digested by TEV protease can specifically bind to mouse anti-GⅡ.17 VP1 polyclonal antibody and assemble into VLPs (diameter:about 40 nm). [Conclusion] In this study, GⅡ.17 VLPs were prepared with E. coli, which can be used as the candidate GⅡ.17 vaccine.