Characterization of particles formed by the precursor protein VPX of infectious bursal disease virus in insect Hi-5 cells: implication on its proteolytic processing.
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 from 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 microg 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.
Lee
Meng-Shiou
MS
Graduate Institute of Biotechnology, National Chung Hsing University, Taichung, Taiwan 40227, ROC.
Wang
Min-Ying
MY
Tai
Yu-Jui
YJ
Lai
Su-Yuan
SY
D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis / Q000276:immunology; D002460:Cell Line; D002645:Chickens; D007243:Infectious bursal disease virus / Q000276:immunology* / Q000378:metabolism / Q000648:ultrastructure; D009500:Neutralization Tests; D011498:Protein Precursors / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013045:Species Specificity; D022223:Vaccines, Subunit / Q000276:immunology; D015678:Viral Structural Proteins / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D014765:Viral Vaccines / Q000276:immunology*
D000914:Antibodies, Viral; D011498:Protein Precursors; D011994:Recombinant Proteins; C082513:VP2 protein, infectious bursal disease virus; D022223:Vaccines, Subunit; D015678:Viral Structural Proteins; D014765:Viral Vaccines