Functional processing and secretion of Chikungunya virus E1 and E2 glycoproteins in insect cells
Autor(es): Metz Stefan W, Geertsema Corinne, Martina Byron E, Andrade Paulina, Heldens Jacco G, van Oers Monique M, Goldbach Rob W, Vlak Just M, Pijlman Gorben P
Resumo: Chikungunya virus (CHIKV) is a mosquito-borne, arthrogenic Alphavirus that causes large epidemics in Africa, South-East Asia - India. Recently, CHIKV has been transmitted to humans in Southern Europe by invading - now established Asian tiger mosquitoes. To study the processing of envelope proteins E1 - E2 - to develop a CHIKV subunit vaccine, C-terminally his-tagged E1 - E2 envelope glycoproteins were produced at high levels in insect cells with baculovirus vectors using their native signal peptides located in CHIKV 6K - E3, respectively. Expression in the presence of either tunicamycin or furin inhibitor showed that a substantial portion of recombinant intracellular E1 - precursor E3E2 was glycosylated, but that a smaller fraction of E3E2 was processed by furin into mature E3 - E2. Deletion of the C-terminal transmembrane domains of E1 - E2 enabled secretion of furin-cleaved, fully processed E1 - E2 subunits, which could then be efficiently purified from cell culture fluid via metal affinity chromatography. Confocal laser scanning microscopy on living baculovirus-infected Sf21 cells revealed that full-length E1 - E2 translocated to the plasma membrane, suggesting similar posttranslational processing of E1 - E2, as in a natural CHIKV infection. Baculovirus-directed expression of E1 displayed fusogenic activity as concluded from syncytia formation. CHIKV-E2 was able to induce neutralizing antibodies in rabbits. Chikungunya virus glycoproteins could be functionally expressed at high levels in insect cells - are properly glycosylated - cleaved by furin. The ability of purified, secreted CHIKV-E2 to induce neutralizing antibodies in rabbits underscores the potential use of E2 in a subunit vaccine to prevent CHIKV infections.
Imprenta: Virology Journal, v. 8, p. 353, 2011
Identificador do objeto digital: 10.1186/1743-422X-8-353
Descritores: Chikungunya virus - Biosynthesis ; Chikungunya virus - Cell ; Chikungunya virus - Pathogenesis ; Chikungunya virus - Protein synthesis ; Chikungunya virus - Proteins ; Chikungunya virus - Antibodies ; Chikungunya virus - Viral infections ; Chikungunya Virus - Virus ; Chikungunya virus - Vaccine ; Chikungunya virus - Epidemic
Data de publicação: 2011
