title: Secretory expression of Porcine Circovirus Type 2 capsid protein in Pichia pastoris.
abstract: Porcine circovirus type 2 (PCV2) is associated with postweaning multisystemic wasting syndrome (PMWS). The PCV2 capsid (Cap) protein is a leading antigen candidate for vaccine and serological diagnostic testing, due to its immunogenic properties. In this study, the codon-optimized PCV2 Cap gene was cloned into a pPICZαA vector for secretory expression in the methylotrophic yeast Pichia pastoris after methanol induction. The screening of recombinant yeasts was followed by detection of the recombinant Cap (rCap) protein by Western blot, using sera from pigs naturally infected with PCV2. The rCap secreted protein was used without prior purification as a coating antigen in the ELISA test, with high discrimination between PCV2-positive and negative sera. These results reveal a high confidence in the specific immunoreactivity of the secreted antigen and show the antigenicity of the recombinant protein. The feasibility of the P. pastoris expression system for the production of PCV2 Cap as secreted protein and its apparent bioactivity, suggests there are good prospects for the use of this antigen in the investigation of PCV2 infections and testing for vaccine purposes.
authors: Department of Genetics, Federal University of Pernambuco, Recife, Pernambuco, Brazil.
authors: Jackeline G
authors: JG
authors: Silva
authors: Department of Genetics, Federal University of Pernambuco, Recife, Pernambuco, Brazil.
authors: Eliane C
authors: EC
authors: Coimbra
authors: Department of Genetics, Federal University of Pernambuco, Recife, Pernambuco, Brazil.
authors: André L S
authors: AL
authors: Jesus
authors: Department of Genetics, Federal University of Pernambuco, Recife, Pernambuco, Brazil.
authors: Filipe C
authors: FC
authors: Mariz
authors: Department of Veterinary Medicine, Federal Rural University of Pernambuco, Recife, Pernambuco, Brazil.
authors: Karen M G
authors: KM
authors: Silva
authors: Department of Preventive Veterinary Medicine, Federal University of Minas Gerais, School of Veterinary Medicine, Minas Gerais, Brazil.
authors: Zélia I P
authors: ZI
authors: Lobato
authors: Department of Veterinary Medicine, Federal Rural University of Pernambuco, Recife, Pernambuco, Brazil.
authors: Ana C
authors: AC
authors: Campos
authors: Department of Veterinary Medicine, Federal Rural University of Pernambuco, Recife, Pernambuco, Brazil.
authors: Luciana C A
authors: LC
authors: Coutinho
authors: Department of Veterinary Medicine, Federal Rural University of Pernambuco, Recife, Pernambuco, Brazil.
authors: Roberto S
authors: RS
authors: Castro
authors: Department of Genetics, Federal University of Pernambuco, Recife, Pernambuco, Brazil. Electronic address: antonio.freitas@pq.cnpq.br.
authors: Antonio C
authors: AC
authors: Freitas
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D000956:Antigens, Viral / Q000235:genetics; D015153:Blotting, Western; D036022:Capsid Proteins / Q000235:genetics* / Q000378:metabolism*; D017925:Circovirus / Q000235:genetics*; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D004718:Saccharomycetales; D017422:Sequence Analysis, DNA; D013552:Swine
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D036022:Capsid Proteins; D004279:DNA, Viral; D011994:Recombinant Proteins
keywords: ELISA; PMWS; Pichia pastoris; Porcine circovirus