Epitope mapping of the nucleocapsid protein of European and North American isolates of porcine reproductive and respiratory syndrome virus.
Two major genotypes of porcine reproductive and respiratory syndrome virus (PRRSV) have been described, which correspond to the European and North American isolates. PRRSV nucleocapsid (N) protein has been identified as the most immunodominant viral protein. The N genes from two PRRSV isolates, Olot/91 (European) and Québec 807/94 (North American), were cloned and expressed in: (i) baculovirus under the control of the polyhedrin promoter and (ii) Escherichia coli using the pET3x system. The N protein from both isolates was expressed much more efficiently in E. coli as a fusion protein than in baculovirus. The antigenicity of the protein was similar in both systems and it was recognized by a collection of 48 PRRSV-positive pig sera. The antigenic structure of the PRRSV N protein was investigated using seven monoclonal antibodies (MAbs) and overlapping fragments of the protein expressed in E. coli. Four MAbs recognized two discontinuous epitopes that were present in the partially folded protein, or at least a large fragment comprising the first 78 residues. The other three MAbs revealed the presence of a common antigenic site localized in the central region of the protein (amino acids 50-66). This region is well conserved among different isolates of European and North American origin and is the most hydrophilic region of the protein. However, this epitope, although recognized by the MAbs and many pig sera, is not useful for diagnostic purposes. Moreover, none of the N protein fragments were able to mimic the antigenicity of the entire protein.
Rodriguez
M J
MJ
INGENASA, Madrid, Spain.
Sarraseca
J
J
Garcia
J
J
Sanz
A
A
Plana-Durán
J
J
Ignacio Casal
J
J
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D003001:Cloning, Molecular; D018604:Epitope Mapping; D004926:Escherichia coli / Q000235:genetics; D005060:Europe; D005822:Genetic Vectors; D007106:Immune Sera; D016056:Immunodominant Epitopes / Q000032:analysis / Q000737:chemistry; D008969:Molecular Sequence Data; D019251:Nucleocapsid / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D017924:Nucleopolyhedroviruses / Q000235:genetics; D016366:Open Reading Frames / Q000235:genetics; D019316:Porcine respiratory and reproductive syndrome virus / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D011487:Protein Conformation; D011792:Quebec; D011993:Recombinant Fusion Proteins; D017422:Sequence Analysis, DNA; D013552:Swine
D000911:Antibodies, Monoclonal; D007106:Immune Sera; D016056:Immunodominant Epitopes; D011993:Recombinant Fusion Proteins