title: Mutant canine oral papillomavirus L1 capsid proteins which form virus-like particles but lack native conformational epitopes.
abstract: Recently, the L1 capsid protein of canine oral papillomavirus (COPV) has been used as an effective systemic vaccine that prevents viral infections of the oral mucosa. The efficacy of this vaccine is critically dependent upon native L1 conformation and, when purified from Sf9 insect cells, the L1 protein not only displays type-specific, conformation-dependent epitopes but it also assembles spontaneously into virus-like particles (VLPs). To determine whether VLP formation was coupled to the expression of conformation-dependent epitopes, we generated a series of N- and C-terminal L1 deletion mutants and evaluated their ability to form VLPs (by electron microscopy) and to react with conformation-dependent antibodies (by immunofluorescence microscopy). We found that (a) deletion of the 26 C-terminal residues generated a mutant protein which formed VLPs efficiently and folded correctly both in the cytoplasm and in the nucleus; (b) further truncation of the L1 C terminus (67 amino acids) resulted in a capsid protein which formed VLPs but which failed to express conformational epitopes; (c) deletion of the first 25 N-terminal amino acids also abolished expression of conformational epitopes (without altering VLP formation) but the native conformation of this deletion mutant could be restored by the addition of the human papillomavirus type 11 N terminus. These results demonstrate that VLP formation and conformational epitope expression can be dissociated and that the L1 N terminus has a critical role in protein folding. In addition, it appears that correct L1 protein folding is not dependent upon the nucleoplasmic environment.
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC 20007, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Chen
authors.forename: S J
authors.initials: SJ
authors.lastname: Ghim
authors.forename: A B
authors.initials: AB
authors.lastname: Jenson
authors.forename: R
authors.initials: R
authors.lastname: Schlegel
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000914:Antibodies, Viral; D001483:Base Sequence; D002213:Capsid / Q000737:chemistry* / Q000235:genetics* / Q000276:immunology; D036022:Capsid Proteins; D017931:DNA Primers / Q000235:genetics; D004285:Dogs; D000939:Epitopes / Q000737:chemistry / Q000235:genetics; D006801:Humans; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009154:Mutation; D027383:Papillomaviridae / Q000235:genetics* / Q000276:immunology / Q000502:physiology; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D017510:Protein Folding; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D017384:Sequence Deletion; D014614:Vaccines, Synthetic / Q000737:chemistry / Q000235:genetics; D014765:Viral Vaccines / Q000737:chemistry / Q000235:genetics
chemical_list: D000914:Antibodies, Viral; D036022:Capsid Proteins; D017931:DNA Primers; D000939:Epitopes; C097166:L1 protein, canine oral papillomavirus; D011994:Recombinant Proteins; D014614:Vaccines, Synthetic; D014765:Viral Vaccines