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標題: Characterization of particles formed by the precursor protein VPX of infectious bursal disease virus in insect Hi-5 cells: implication on its proteolytic processing
作者: Lee, M.S.
Wang, M.Y.
Tai, Y.J.
Lai, S.Y.
關鍵字: infectious bursal disease virus;virus-like particle;baculovirus;expression system;immobilized metal-ion affinity chromatography;gel-filtration chromatograph;ion affinity-chromatography;dependent rna-polymerase;capsid protein;maturation process;escherichia-coli;genomic segment;cleavage sites;expression;identification;purification
Project: Journal of Virological Methods
期刊/報告no:: Journal of Virological Methods, Volume 121, Issue 2, Page(s) 191-199.
The precursor (VPX) of host immunogen VP2 protein for infectious bursal disease virus (IBDV) was expressed in insect Sf9 and Hi-5 cells, and the types of particles generated as well as the immunogenicity induced by these particles were examined. Recombinant VPXH (rVPXH) protein, expressed in Hi-5 cells at an expression level 4x higher than in Sf9 cells, was efficiently processed by proteases to yield VP2-like proteins with corresponding molecular weight, a phenomenon not observed previously. At least three structures of particles were observed for VPXH and VP2-like proteins purified by immobilized metal-ion affinity chromatography (MAC). In addition to the two previously identified twisted tubular and isometric particle structures, there was a new one: icosahedral particles of approximately 25 nm in diameter. The purified particles were further separated by gel-filtration chromatography (GFC) linking with HPLC, which was able to resolve the isometric front icosahedral particles better than ultracentrifugation. Chromatographic results indicate that rVPXH protein mainly involved in the formation of the isometric particle structure and occasionally twisted tubular structure, and the icosahedral particles were formed by the degraded products of rVPXH (VP2-like proteins). Thus, by combining IMAC and GFC, it was shown that VPX was processed efficiently to yield VP2-like protein that could form small virus-like particles in Hi-5 cells. Finally, we demonstrated that virus-neutralizing antibodies were induced when susceptible chickens were vaccinated with the IMAC-purified rVPXH protein (40 mug per bird). This indicates that these particles are highly immunogenic and might serve as an alternative vaccine candidate for the development of IBDV subunit vaccine. (C) 2004 Elsevier B.V. All rights reserved.
ISSN: 0166-0934
DOI: 10.1016/j.viromet.2004.06.014
Appears in Collections:生物科技學研究所

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