A plant-based transient expression system for the rapid production of highly immunogenic Hepatitis E virus-like particles.

OBJECTIVE: Hepatitis E virus (HEV) infection is a major cause of acute hepatitis worldwide. The aim of the study is the development of plant expression system for the production of virus-like particles formed by HEV capsid and the characterization of their immunogenicity. RESULTS: Open reading frame (ORF) 2 encodes the viral capsid protein and possesses candidate for vaccine production. In this study, we used truncated genotype 3 HEV ORF 2 consisting of aa residues 110 to 610. The recombinant protein was expressed in Nicotiana benthamiana plants using the self-replicating potato virus X-based vector pEff up to 10% of the soluble protein fraction. The yield of HEV 110-610 after purification was 150-200 µg per 1 g of green leaf biomass. The recombinant protein formed nanosized virus-like particles. The immunization of mice with plant-produced HEV 110-610 protein induced high levels of HEV-specific serum antibodies. CONCLUSIONS: HEV ORF 2 (110-610 aa) can be used as candidate for the development of a plant-produced vaccine against Hepatitis E.

Rapid High-Yield Transient Expression of Swine Hepatitis E ORF2 Capsid Proteins in Nicotiana benthamiana Plants and Production of Chimeric Hepatitis E Virus-Like Particles Bearing the M2e Influenza Epitope.

The Hepatitis E virus (HEV) is a causative agent of acute hepatitis, mainly transmitted by the fecal-oral route or zoonotic. Open reading frame (ORF) 2 encodes the viral capsid protein, which is essential for virion assembly, host interaction, and inducing neutralizing antibodies. In this study, we investigated whether full-length and N- and C-terminally modified versions of the capsid protein transiently expressed in N. benthamiana plants could assemble into highly-immunogenic, virus-like particles (VLPs). We also assessed whether such VLPs can act as a carrier of foreign immunogenic epitopes, such as the highly-conserved M2e peptide from the Influenza virus. Plant codon-optimized HEV ORF2 capsid genes were constructed in which the nucleotides coding the N-terminal, the C-terminal, or both parts of the protein were deleted. The M2e peptide was inserted into the P2 loop after the residue Gly556 of HEV ORF2 protein by gene fusion, and three different chimeric constructs were designed. Plants expressed all versions of the HEV capsid protein up to 10% of total soluble protein (TSP), including the chimeras, but only the capsid protein consisting of aa residues 110 to 610 (HEV 110-610) and chimeric M2 HEV 110-610 spontaneously assembled in higher order structures. The chimeric VLPs assembled into particles with 22-36 nm in diameter and specifically reacted with the anti-M2e antibody.