Folding of the rabbit hemorrhagic disease virus capsid protein and delineation of N-terminal domains dispensable for assembly.
Rabbit hemorrhagic disease virus (RHDV) and European brown hare syndrome virus (EBHSV) are caliciviruses that produce severe symptoms and are lethal to rabbits and hares. The folding of the capsid protein was studied by determination of the antigenic pattern of chimeric capsid proteins, composed of regions from RHDV and EBHSV capsid proteins. The anti-RHDV monoclonal antibody (MAb) E3, which is known to bind an external conformational epitope, recognized the RHDV C-terminal region. The anti-RHDV MAb A47, which binds a buried epitope, recognized the RHDV N-terminal region. Using a pGEX expression library, we more precisely mapped the MAb A47 epitope on a 31 residues length peptide, between residue 129 and 160 of the VP60, confirming its location in the N-terminal part of the protein. These results demonstrate that the C-terminal part of the protein is accessible to the exterior whereas the N-terminal domain of the protein constitutes the internal shell domain of the particle. With the aim of using virus-like particles (VLPs) of RHDV as epitope carriers or DNA transfer vectors, we produced in the baculovirus system three proteins, DeltaN1, DeltaN2 and DeltaN3, truncated at the N terminus. The DeltaN1 protein assembled into VLPs, demonstrating that the first 42 amino acid residues are not essential for capsid assembly. In contrast, DeltaN2, from which the first 75 residues were missing, was unable to form VLPs. The small particles obtained with the DeltaN3 protein lacking residues 31 to 93, located in the immunodominant region of the RHDV capsid protein, indicate that up to 62 amino acid residues can be eliminated without preventing assembly.
Laboratoire de Virologie et Barrière d'Espèce, UR 086, INRA, Centre de recherches de Tours, F-37380 Nouzilly, France. slaurent@tours.inra.fr
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Laurent
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Kut
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Remy-Delaunay
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Rasschaert
D000595:Amino Acid Sequence; D000818:Animals; D002213:Capsid / Q000737:chemistry*; D018604:Epitope Mapping; D017929:Hemorrhagic Disease Virus, Rabbit / Q000737:chemistry* / Q000502:physiology; D008969:Molecular Sequence Data; D017510:Protein Folding; D011817:Rabbits; D018411:Spodoptera; D014771:Virion / Q000502:physiology; D019065:Virus Assembly