Self-assembly of the infectious bursal disease virus capsid protein, rVP2,expressed in insect cells and purification of immunogenic chimeric rVP2H particles by immobilized metal-ion affinity chromatography

A gene encoding a structural protein (VP2) of a local strain (P3009) of inf ectious bursal disease virus (IBDV) was cloned and expressed using the bacu lovirus expression system to develop a subunit vaccine against IBDV infecti on in Taiwan. The expressed rVP2 proteins formed particles of approximately 20-30 nm in diameter. Those particles were partially purified employing su crose density gradient ultracentrifugation, and the purified particles were recognized by a monoclonal antibody against the VP2 protein of IBDV P3009. To facilitate the purification of the particles, the VP2 protein was engin eered to incorporate a metal ion binding site (His)(6) at its C-terminus. T he chimeric rVP2H proteins also formed particles, which could be affinity;p urified in one step with immobilized metal ions (Ni2+). Particle formation was confirmed by direct observation under the electron microscope. The prod uction level of rVP2H protein was determined to be 20 mg/L in a batch cultu re of Hi-5 cells by quantifying the concentration of the purified proteins. The chicken protection assay was performed to evaluate the immunogenicity of the rVP2H protein. When susceptible chickens were inoculated with the re combinant rVP2H proteins (40 mu g/bird), virus-neutralizing antibodies were induced, thereby conferring a high level of protection against the challen ge of a very virulent strain of IBDV. In conclusion, the most significant f inding in this work is that both of the expressed rVP2 and rVP2H proteins c an form a particulate structure capable of inducing a strong immunological response in a vaccinated chicken. (C) 2000 John Wiley at Sons, Inc.