MUTANT CANINE ORAL PAPILLOMAVIRUS L1 CAPSID PROTEINS WHICH FORM VIRUS-LIKE PARTICLES BUT LACK NATIVE CONFORMATIONAL EPITOPES

Recently, the L1 capsid protein of canine oral papillomavirus (COPV) h as been used as an effective systemic vaccine that prevents viral infe ctions of the oral mucosa, The efficacy of this vaccine is critically dependent upon native L1 conformation and, when purified from Sf9 inse ct cells, the L1 protein not only displays type-specific, conformation -dependent epitopes but it also assembles spontaneously into virus-lik e particles (VLPs), To determine whether VLP formation was coupled to the expression of conformation-dependent epitopes, we generated a seri es of N- and C-terminal L1 deletion mutants and evaluated their abilit y to form VLPs (by electron microscopy) and to react with conformation -dependent antibodies (by immunofluorescence microscopy). We found tha t (a) deletion of the 26 C-terminal residues generated a mutant protei n which formed VLPs efficiently and folded correctly both in the cytop lasm and in the nucleus; (b) further truncation of the L1 C terminus ( 67 amino acids) resulted in a capsid protein which formed VLPs but whi ch failed to express conformational epitopes; (c) deletion of the firs t 25 N-terminal amino acids also abolished expression of conformationa l epitopes (without altering VLP formation) but the native conformatio n of this deletion mutant could be restored by the addition of the hum an papillomavirus type 11 N terminus. These results demonstrate that V LP 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.