title: Inactivation and purification of cowpea mosaic virus-like particles displaying peptide antigens from Bacillus anthracis.
abstract: Chimeric cowpea mosaic virus (CPMV) particles displaying foreign peptide antigens on the particle surface are suitable for development of peptide-based vaccines. However, commonly used PEG precipitation-based purification methods are not sufficient for production of high quality vaccine candidates because they do not allow for separation of chimeric particles from cleaved contaminating species. Moreover, the purified particles remain infectious to plants. To advance the CPMV technology further, it is necessary to develop efficient and scalable purification strategies and preferably eliminate the infectivity of chimeric viruses. CPMV was engineered to display a 25 amino acid peptide derived from the Bacillus anthracis protective antigen on the surface loop of the large coat protein subunit. The engineered virus was propagated in cowpea plants and assembled into chimeric virus particles displaying 60 copies of the peptide on the surface. An effective inactivation method was developed to produce non-infectious chimeric CPMV virus-like particles (VLPs). Uncleaved VLPs were separated from the contaminating cleaved forms by anion exchange chromatography. The yield of purified chimeric VLPs was 0.3 g kg(-1) of leaf tissue. The results demonstrate the ability to generate multi-gram quantities of non-infectious, chimeric CPMV VLPs in plants for use in the development of peptide-based vaccines.
authors.affiliation: The Dow Chemical Company, Dowpharma, San Diego, CA 92121, USA. PhelpsJ@colorado.edu <PhelpsJ@colorado.edu>
authors.forename: Jamie P
authors.initials: JP
authors.lastname: Phelps
authors.forename: Nghiep
authors.initials: N
authors.lastname: Dang
authors.forename: Lada
authors.initials: L
authors.lastname: Rasochova
mesh_terms: D000645:Ammonium Sulfate; D000942:Antigens, Bacterial / Q000235:genetics / Q000378:metabolism; D001408:Bacillus anthracis / Q000276:immunology; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics; D002852:Chromatography, Ion Exchange; D017798:Comovirus / Q000254:growth & development / Q000302:isolation & purification* / Q000378:metabolism; D007887:Fabaceae / Q000821:virology; D006863:Hydrogen-Ion Concentration; D007218:Industrial Microbiology / Q000379:methods*; D018515:Plant Leaves / Q000821:virology; D016865:Reassortant Viruses / Q000254:growth & development / Q000302:isolation & purification* / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis; D038301:Virus Inactivation
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D011994:Recombinant Proteins; D000645:Ammonium Sulfate