978. Production of SARS Coronavirus-Like Particles That Bind Host Cells and Serve as Vaccine Antigen

Abstract

Top of pageAbstract Severe Acute Respiratory Syndrome (SARS) is a deadly form of pneumonia caused by a recently identified coronavirus (SARS-CoV). Viral particles of SARS-CoV comprise three viron structural proteins, including spike (S), membrane (M), and envelope (E). To efficiently express S, M, and E proteins of SARS-CoV simultaneously in one cell, we constructed a plasmid harboring the three genes with GFP (green fluorescence protein) fusion to the M protein. Expression of the three genes is engineered under regulation of tet operon. Stable transfection of this plasmid into Vero E6 cells creates cell lines showing inducible expression of virus-like particles (VLPs). Expression and packaging of VLP in cytoplasm is observed by confocal microscopy. We then purified the secreted VLPs from cell culture medium through ultra-centrifugation. Containment of each protein in VLPs was assured by coomassie blue staining and western blotting. Interestingly, two types of particles were secreted by one of our producing cell lines. One contains M dominantly, the other contains S dominantly. Electron microscopy reveals homogenous particles of both VLPs. We have further demonstrated binding capability of M dominant VLP to Vero E6 cells as host by flow cytometry and confocal microscopy analyses. Immunization of the two VLPs induced antibodies against both VLPs in ELISA and western blot, suggesting a promising immunogenicity of both VLPs as vaccine antigen. The genetic engineered VLPs bearing resemblance to the authentic SARS-CoV as well as their antibodies are important tools toward development of effective vaccine against highly contingent infectious disease like SARS.