Modified capsid proteins containing at least a portion of hepatitis E virus (HEV) open reading frame 2 (ORF2) having one or more cysteine residues in a surface variable loop or the C-terminus of HEV ORF2, or a portion thereof, are provided. The modified capsid proteins can be used to form hepatitis E virus (HEV) virus like particles (VLPs) having cysteine functional groups exposed on the outer-surface. The exposed cysteine functional groups can be modified via their thiol reactive group. For example, a bioactive agent, such as a cell-targeting ligand, can be conjugated to the one or more cysteines for targeted delivery of chemically activated nanocapsids.

Genes for proteins which spontaneously form dimers and/or oligomers can be recombinantly linked together, which upon expression in E. coli produces stable dimeric fusion proteins that spontaneously self-assemble into enormous, polyvalent complexes having increased immunogenicity and functionality. Linear, network and agglomerate complexes with enormous sizes and polyvalences are constructed using glutathione S-transferase, Norovirus P domains (NoV P.sup. and NoV.sup.+), the protruding (P) domain of hepatitis E virus (HEV P), the astrovirus P domain (AstV), a monomeric peptide epitope (M2e of influenza virus), and/or a protein antigen (VP8* of rotavirus) fused in different combinations. The resulting complexes can contain hundreds to thousands NoV P-protein, HEV, AstV, M2e and/or VP8* copies and exhibit higher immunogenicity than the individual proteins alone. The large size and multivalent nature of the complexes are candidates as a bivalent or multivalent vaccines against Norovirus and other pathogens, and for generation of antibodies for diagnosis and research purposes.

Provided in the present invention are a diphtheria toxin non-toxic mutant CRM197 or a fragment thereof as an adjuvant in a fusion protein and the use thereof to enhance the immunogenicity of a target protein fused therewith, for example, an HEV capsid protein, or an influenza virus M2 protein or an immunogenic fragment thereof. Also provided is a method for enhancing the immunogenicity of a target protein, comprising the fusion expression of the CRM197 or the fragment thereof with the target protein to form a fusion protein. Further provided is a fusion protein comprising the CRM197 or the fragment thereof and a target protein, the CRM197 or the fragment thereof enhancing the immunogenicity of the target protein. The present invention also provides an isolated nucleic acid encoding the fusion protein, a construct and a vector comprising said nucleic acid, and a host cell comprising the nucleic acid.

Provided in the present invention are a diphtheria toxin non-toxic mutant CRM197 or a fragment thereof as an adjuvant in a fusion protein and the use thereof to enhance the immunogenicity of a target protein fused therewith, for example, an HEV capsid protein, or an influenza virus M2 protein or an immunogenic fragment thereof. Also provided is a method for enhancing the immunogenicity of a target protein, comprising the fusion expression of the CRM197 or the fragment thereof with the target protein to form a fusion protein. Further provided is a fusion protein comprising the CRM197 or the fragment thereof and a target protein, the CRM197 or the fragment thereof enhancing the immunogenicity of the target protein. The present invention also provides an isolated nucleic acid encoding the fusion protein, a construct and a vector comprising said nucleic acid, and a host cell comprising the nucleic acid.

The present disclosure provides affinity tagged heterodimeric polypeptides comprising a hepatitis C virus (HCV) E1 polypeptide and an HCV E2 polypeptide, where one or both of the E1 and E2 polypeptides comprises an affinity tag. The present disclosure provides a method of producing an affinity tagged E1/E2 heterodimer of the present disclosure. The present disclosure provides methods of producing untagged HCV E1/E2 heterodimers. The present disclosure provides HCV E1/E2 heterodimers, compositions comprising same, and methods of inducing an immune response to HCV.