The effect of C-terminal end of the Infectious Bursal Disease Virus
Structural Protein, VP2, on its Self-Assembly in Insect Cells
Min-Ying Wang1*, Meng-Shiou Lee1, Shyue-Ru Doong3, Ji-Yi Ho1 and Long-Huw
Lee2
1
3
Graduate Institute of Agricultural Biotechnology, and 2Department of Veterinary
Medicine National Chung Hsing University, Taichung, Taiwan 40227 ROC
Department of Chemical Engineering, TangHai University, 181 Taichungkang Rd.,
Sec. 3, Taichung, Taiwan, ROC
*To whom correspondence should be addressed
Mailing Address: Graduate Institute of Agricultural Biotechnology,
National Chung Hsing University, Taichung, 40227 Taiwan
Telephone: 886-4-285-6697
Fax: 886-4-285-6697, 886-4-285-3527
E-mail address: mywang@dragon.nchu.edu.tw
Running title: Self-Assembly of IBDV rVP2 Proteins and Purification of the
Immunogenic rVP2H Particles.
Abstract
IBDV is the member of the Avibirnavirus genus in the Birnaviridae family and its
genome is formed by two double-stranded RNA segments.
VP2 is one of the major
structure proteins of IBDV capsids and the primary host-protective immunogen of
IBDV.
When VP2 fused with an extra six-hitidine-residue at the C-terminus
(rVP2H) was expressed alone in insect cells, highly immunogenic virus-like particles
formed spontaneously.
To understand the criteria for the formation of the rVP2H
particles, the effect of C-terminal region of VPX on the capability to form the
particulate structure was investigated in this report.
Studies were performed with
recombinant baculoviruses expressed in Sf9 cells and Hi-5 cells. To facilitate the
purification of the particles, the recombinant mutant proteins were incorporated a
metal ion binding site (His)6 at their C-terminus. Experimental data indicated that two
shorter versions of rVP2H, one contract 15 amino acids (rVPXD15H) and the other
(rVP2D41H) 41 from the VP2 core sequence 1M-R452, also formed the particulate
structure. The morphologies of rVP2D15 and rVP2D41 particles were similar to
that formed by rVP2H particles, dodecahedral particles with a diameter,
approximately, 23 nm in size. Deletion of 53 or 63 amino acids from the C-terminal
end of rVP2H abolishes the formation of particles, suggesting the C-terminal regions,
amino acid residues ranging from 399 to 411, is critical for the particulate formation.
Key words: infectious bursal disease virus; virus-like particle; baculovirus expression
system; immobilized metal-ion affinity chromatography.