A broadly cross-reactive monoclonal antibody against hepatitis E virus capsid antigen

Abstract

To generate a hepatitis E virus (HEV) genotype 3 (HEV-3)–specific monoclonal antibody (mAb), the Escherichia coli–expressed carboxy-terminal part of its capsid protein was used to immunise BALB/c mice. The immunisation resulted in the induction of HEV-specific antibodies of high titre. The mAb G117-AA4 of IgG1 isotype was obtained showing a strong reactivity with the homologous E. coli, but also yeast-expressed capsid protein of HEV-3. The mAb strongly cross-reacted with ratHEV capsid protein derivatives produced in both expression systems and weaker with an E. coli–expressed batHEV capsid protein fragment. In addition, the mAb reacted with capsid protein derivatives of genotypes HEV-2 and HEV-4 and common vole hepatitis E virus (cvHEV), produced by the cell-free synthesis in Chinese hamster ovary (CHO) and Spodoptera frugiperda (Sf21) cell lysates. Western blot and line blot reactivity of the mAb with capsid protein derivatives of HEV-1 to HEV-4, cvHEV, ratHEV and batHEV suggested a linear epitope. Use of truncated derivatives of ratHEV capsid protein in ELISA, Western blot, and a Pepscan analysis allowed to map the epitope within a partially surface-exposed region with the amino acid sequence LYTSV. The mAb was also shown to bind to human patient–derived HEV-3 from infected cell culture and to hare HEV-3 and camel HEV-7 capsid proteins from transfected cells by immunofluorescence assay. The novel mAb may serve as a useful tool for further investigations on the pathogenesis of HEV infections and might be used for diagnostic purposes. Key points • The antibody showed cross-reactivity with capsid proteins of different hepeviruses. • The linear epitope of the antibody was mapped in a partially surface-exposed region. • The antibody detected native HEV-3 antigen in infected mammalian cells.

Serological and molecular survey of hepatitis E virus in cats and dogs in Spain

“…Whenever possible, the presence of antibodies against the capsid proteins of HEV‐A and HEV‐C was assessed in the seropositive samples by western blot analysis as previously described (Kubickova et al., 2021). For this purpose, carboxy‐terminal segments of the capsid proteins of rat HEV‐C1, HEV‐3 and a nucleocapsid protein derivative (amino acid residues 1‐39/213‐433) of Puumala orthohantavirus strain Vranica/Hällnäs, as negative control, were produced as His‐tagged recombinant proteins in Escherichia coli and purified by nickel‐chelate affinity chromatography (Dremsek et al., 2012; Lundkvist et al., 2002).…”

Hepatitis E virus infection activates NOD‐like receptor family pyrin domain‐containing 3 inflammasome antagonizing interferon response but therapeutically targetable

“…Finally, we used purified recombinant ORF2 protein produced by yeast. [ 17 ] Incubation of THP‐1 macrophages with ORF2 protein effectively triggered IL‐1β mRNA expression (Figure S3 G) and IL‐1β protein secretion (Figure 3L ). These results collectively indicate that the ORF2 capsid protein and the integral HEV particle in particular are robust activators of inflammasome response.…”

The Potential of Eukaryotic Cell-Free Systems as a Rapid Response to Novel Zoonotic Pathogens: Analysis of SARS-CoV-2 Viral Proteins

“…A virus like particle derived from the norovirus ( Sheng et al, 2017 ) and Qβ phage ( Smith et al, 2012 ) were synthesized in an E. coli based cell-free system. Using eukaryotic cell-free systems the capsid protein ( Spearman and Ratner, 1996 ) and the envelope protein ( Mikami et al, 2006 ) of the human immunodeficiency virus, the gp67 envelope protein ( Merk et al, 2015 ), a virus like particle from the human papillomavirus ( Wang et al, 2008 ) and the capsid protein of hepatitis E virus ( Kubickova et al, 2021 ) have been synthesized and evaluated. These data show that cell-free protein synthesis can help to characterize single viral proteins and this knowledge can be used to further elucidate the complete virus.…”