Characterization of self-assembled virus-like particles of ferret hepatitis E virus generated by recombinant baculoviruses

Ferret hepatitis E virus (HEV), a novel hepatitis E-like virus, has been identified in ferrets in The Netherlands. Due to the lack of a cell-culture system for ferret HEV, the antigenicity, pathogenicity and epidemiology of this virus have remained unclear. In the present study, we used a recombinant baculovirus expression system to express the 112-N-terminus and 47-C-terminus-amino-acid-truncated ferret HEV ORF2 protein in insect Tn5 cells, and found that a large amount of a 53 kDa protein (F-p53) was expressed and efficiently released into the supernatant. Electron microscopic analysis revealed that F-p53 was self-assembled into virus-like particles (ferret HEV-LPs). These ferret HEV-LPs were estimated to be 24 nm in diameter, which is similar to the size of G1, G3, G4 and rat HEV-LPs derived from both the N-terminus- and C-terminus-truncated constructs. Antigenic analysis demonstrated that ferret HEV-LPs were cross-reactive with G1, G3, G4 and rat HEVs, and rat HEV and ferret HEV showed a stronger cross-reactivity to each other than either did to human HEV genotypes. However, the antibody against ferret HEV-LPs does not neutralize G3 HEV, suggesting that the serotypes of these two HEVs are different. An ELISA for detection of anti-ferret HEV IgG and IgM antibodies was established using ferret HEV-LPs as antigen, and this assay system will be useful for monitoring ferret HEV infection in ferrets as well as other animals. In addition, analysis of ferret HEV RNA detected in ferret sera collected from a breeding colony in the USA revealed the genetic diversity of ferret HEV.

Download Full-text

Characterization of self-assembled virus-like particles of rat hepatitis E virus generated by recombinant baculoviruses

Journal of General Virology ◽

10.1099/vir.0.034835-0 ◽

2011 ◽

Vol 92 (12) ◽

pp. 2830-2837 ◽

Cited By ~ 48

Author(s):

Tian-Cheng Li ◽

Kumiko Yoshimatsu ◽

Shumpei P. Yasuda ◽

Jiro Arikawa ◽

Takaaki Koma ◽

...

Keyword(s):

Hepatitis E Virus ◽

Expression System ◽

Recombinant Baculovirus ◽

Baculovirus Expression System ◽

Hepatitis E ◽

Serum Samples ◽

Electron Microscopic ◽

Rat Serum ◽

Wild Rats ◽

Self Assembled

Hepatitis E virus (HEV) is a causative agent of hepatitis E. Recently, a novel hepatitis E-like virus was isolated from Norway rats in Germany. However, the antigenicity, pathogenicity and epidemiology of this virus are unclear because of the lack of a cell-culture system in which to grow it. In this study, an N-terminally truncated ORF2 protein was expressed in insect Tn5 cells using a recombinant baculovirus expression system and a large amount of 53 kDa protein was expressed and efficiently released into the supernatant. Electron microscopic analyses of the purified 53 kDa protein revealed that the protein self-assembled into two types of empty HEV-like particles (rat HEVLPs). The smaller rat HEVLPs were estimated to be 24 nm in diameter, which is similar to the size of genotype G1, G3 and G4 HEVLPs. The larger rat HEVLPs were estimated to measure 35 nm in diameter, which is similar to the size of native rat HEV particles. An ELISA to detect antibodies was established using rat HEVLPs as the antigens, which demonstrated that rat HEVLPs were cross-reactive with G1, G3 and G4 HEVs. Detection of IgG and IgM antibodies was performed by examination of 139 serum samples from wild rats trapped in Vietnam, and it was found that 20.9 % (29/139) and 3.6 % (5/139) of the samples were positive for IgG and IgM, respectively. In addition, rat HEV RNA was detected in one rat serum sample that was positive for IgM. These results indicated that rat HEV is widespread and is transmitted among wild rats.

Download Full-text

Essential Elements of the Capsid Protein for Self-Assembly into Empty Virus-Like Particles of Hepatitis E Virus

Journal of Virology ◽

10.1128/jvi.79.20.12999-13006.2005 ◽

2005 ◽

Vol 79 (20) ◽

pp. 12999-13006 ◽

Cited By ~ 82

Author(s):

Tian-Cheng Li ◽

Naokazu Takeda ◽

Tatsuo Miyamura ◽

Yoshiharu Matsuura ◽

Joseph C. Y. Wang ◽

...

Keyword(s):

Capsid Protein ◽

Self Assembly ◽

Hepatitis E Virus ◽

Recombinant Baculovirus ◽

Hepatitis E ◽

Essential Elements ◽

Mucosal Vaccine ◽

Sf9 Cells ◽

Reading Frame ◽

Virus Like Particles

ABSTRACT Hepatitis E virus (HEV) is a noncultivable virus that causes acute liver failure in humans. The virus's major capsid protein is encoded by an open reading frame 2 (ORF2) gene. When the recombinant protein consisting of amino acid (aa) residues 112 to 660 of ORF2 is expressed with a recombinant baculovirus, the protein self-assembles into virus-like particles (VLPs) (T.-C. Li, Y. Yamakawa, K. Suzuki, M. Tatsumi, M. A. Razak, T. Uchida, N. Takeda, and T. Miyamura, J. Virol. 71:7207-7213, 1997). VLPs can be found in the culture medium of infected Tn5 cells but not in that of Sf9 cells, and the major VLPs have lost the C-terminal 52 aa. To investigate the protein requirement for HEV VLP formation, we prepared 14 baculovirus recombinants to express the capsid proteins truncated at the N terminus, the C terminus, or both. The capsid protein consisting of aa residues 112 to 608 formed VLPs in Sf9 cells, suggesting that particle formation is dependent on the modification process of the ORF2 protein. In the present study, electron cryomicroscopy and image processing of VLPs produced in Sf9 and Tn5 cells indicated that they possess the same configurations and structures. Empty VLPs were found in both Tn5 and Sf9 cells infected with the recombinant containing an N-terminal truncation up to aa residue 125 and C-terminal to aa residue 601, demonstrating that the aa residues 126 to 601 are the essential elements required for the initiation of VLP assembly. The recombinant HEV VLPs are potential mucosal vaccine carrier vehicles for the presentation of foreign antigenic epitopes and may also serve as vectors for the delivery of genes to mucosal tissue for DNA vaccination and gene therapy. The results of the present study provide useful information for constructing recombinant HEV VLPs having novel functions.

Download Full-text

Production of capsid proteins of rat hepatitis E virus in Escherichia coli and characterization of self-assembled virus-like particles

Virus Research ◽

10.1016/j.virusres.2021.198483 ◽

2021 ◽

pp. 198483

Author(s):

Tominari Kobayashi ◽

Masaharu Takahashi ◽

Satoshi Ohta ◽

Shigeo Nagashima ◽

Putu Prathiwi Primadharsini ◽

...

Keyword(s):

Escherichia Coli ◽

Hepatitis E Virus ◽

Hepatitis E ◽

Capsid Proteins ◽

Virus Like Particles ◽

Self Assembled

Download Full-text

Characterization of self-assembled virus-like particles of dromedary camel hepatitis e virus generated by recombinant baculoviruses

Virus Research ◽

10.1016/j.virusres.2015.06.022 ◽

2015 ◽

Vol 210 ◽

pp. 8-17 ◽

Cited By ~ 13

Author(s):

Xianfeng Zhou ◽

Michiyo Kataoka ◽

Zheng Liu ◽

Naokazu Takeda ◽

Takaji Wakita ◽

...

Keyword(s):

Hepatitis E Virus ◽

Hepatitis E ◽

Recombinant Baculoviruses ◽

Dromedary Camel ◽

Virus Like Particles ◽

Self Assembled

Download Full-text

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

Biotechnology Letters ◽

10.1007/s10529-020-02995-x ◽

2020 ◽

Vol 42 (11) ◽

pp. 2441-2446

Author(s):

Eugenia S. Mardanova ◽

Katerina H. Takova ◽

Valentina T. Toneva ◽

Gergana G. Zahmanova ◽

Liudmila M. Tsybalova ◽

...

Keyword(s):

Transient Expression ◽

Hepatitis E Virus ◽

Expression System ◽

Hepatitis E ◽

Virus Like Particles ◽

Transient Expression System ◽

Rapid Production

Download Full-text

Efficient Production of Chimeric Hepatitis B Virus-Like Particles Bearing an Epitope of Hepatitis E Virus Capsid by Transient Expression in Nicotiana benthamiana

Life ◽

10.3390/life11010064 ◽

2021 ◽

Vol 11 (1) ◽

pp. 64

Author(s):

Gergana Zahmanova ◽

Milena Mazalovska ◽

Katerina Takova ◽

Valentina Toneva ◽

Ivan Minkov ◽

...

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Nicotiana Benthamiana ◽

Hepatitis E Virus ◽

Chimeric Protein ◽

Hepatitis E ◽

Core Antigen ◽

Efficient Production ◽

Virus Like Particles ◽

B Virus

The core antigen of hepatitis B virus (HBcAg) is capable of self-assembly into virus-like particles (VLPs) when expressed in a number of heterologous systems. Such VLPs are potential carriers of foreign antigenic sequences for vaccine design. In this study, we evaluated the production of chimeric HBcAg VLPs presenting a foreign epitope on their surface, the 551–607 amino acids (aa) immunological epitope of the ORF2 capsid protein of hepatitis E virus. A chimeric construct was made by the insertion of 56 aa into the immunodominant loop of the HBcAg. The sequences encoding the chimera were inserted into the pEAQ-HT vector and infiltrated into Nicotiana benthamiana leaves. The plant-expressed chimeric HBcHEV ORF2 551–607 protein was recognized by an anti-HBcAg mAb and anti-HEV IgG positive swine serum. Electron microscopy showed that plant-produced chimeric protein spontaneously assembled into “knobbly” ~34 nm diameter VLPs. This study shows that HBcAg is a promising carrier platform for the neutralizing epitopes of hepatitis E virus (HEV) and the chimeric HBcAg/HEV VLPs could be a candidate for a bivalent vaccine.

Download Full-text