Intracellular Expression of Hepatitis E Virus ORF2 Protein in pichia pastoris and Its Purification
We have used the methytotrophic yeast pichia pastoris to express the open reading frame 2(ORF2) protein of hepatitis E virus (HEV). The gene which encodes amino acid residues 69～660 of ORF2, was amplified from cDNA of HEV by PCR and cloned into the pichia expression vector pPIC3.5K. The recombinant plasmid was transformed into GS115 and screened for G418 resistant and Mut phenotype. The high resistant Muts clones were chosen to test the expression of HEV ORF2 protein in BMGY/BMMY media by SDS-PAGE and ELISA. The high expressing clone was scaled up and the recombinant products were purified by affinity chromatography. The expressed protein was estimated to be 59kDa with 96% purity by scanning analysis. Western blot showed it could be recognized by the McAb of HEV ORF2. The successful expression of HEV ORF2 protein in p. pastoris and the production of recombinant protein provides basis for genetically engineered vaccine development of hepatitis E.