Expression and self-assembly of empty virus-like particles of hepatitis E virus.
Hepatitis E virus (HEV) is a pathogenic agent that causes fecally-orally transmitted acute hepatitis. The genome, a single-stranded positive-sense RNA, encodes three forward open reading frames (ORFs), in which an approximately 2-kb structural protein is located in the 3' end. To produce HEV-like particles the structural protein, with its N terminus truncated (amino acid residues 112 to 660 of ORF2), was expressed in insect Tn5 cells by a recombinant baculovirus. In addition to the primary translation product with a molecular mass of 58 kDa, a large amount of a further-processed molecule with a molecular mass of 50 kDa was generated and efficiently released into the culture medium. Electron microscopic observation of the culture medium revealed that the 50-kDa protein self-assembled to form empty virus-like particles (VLPs). The buoyant density of the VLPs in CsCl was 1.285 g/cm3 and their diameter was 23.7 nm, a little smaller than the 27 nm of native HEV particles secreted into the bile or stools of experimentally infected monkeys. The yield of the VLPs was 1 mg per 10(7) cells as a purified form. The particles possess antigenicity similar to that of authentic HEV particles and, consequently, they appear to be a good antigen for the sensitive detection of HEV-specific immunoglobulin G (IgG) and IgM antibodies. Furthermore, the VLP may be the most promising candidate yet for an HEV vaccine, owing to its potent immunogenicity.
Li
T C
TC
Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.
Yamakawa
Y
Y
Suzuki
K
K
Tatsumi
M
M
Razak
M A
MA
Uchida
T
T
Takeda
N
N
Miyamura
T
T
D000818:Animals; D016367:Baculoviridae; D001646:Bile / Q000821:virology; D015153:Blotting, Western; D002213:Capsid / Q000096:biosynthesis; D002460:Cell Line; D004797:Enzyme-Linked Immunosorbent Assay; D005243:Feces / Q000821:virology; D016679:Genome, Viral; D006508:Hepatitis Antibodies / Q000097:blood; D016751:Hepatitis E / Q000276:immunology / Q000635:transmission; D016752:Hepatitis E virus / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D006801:Humans; D008253:Macaca mulatta; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000096:biosynthesis; D018411:Spodoptera; D014162:Transfection; D015678:Viral Structural Proteins / Q000096:biosynthesis* / Q000648:ultrastructure
D006508:Hepatitis Antibodies; D011994:Recombinant Proteins; D015678:Viral Structural Proteins