S1 domain of the porcine epidemic diarrhea virus spike protein as a vaccine antigen

Makadiya, Niraj; Brownlie, Robert; van den Hurk, Jan; Berube, Nathalie; Allan, Brenda; Gerdts, Volker; Zakhartchouk, Alexander

Mammalian expression of S1 utilizing a human CMV promoter within an episomal vector was also investigated. In preliminary experiments, expression of S1 in which the native signal peptide was replaced with that of tissue plaminogen activator (TPA) was compared with expression of S1 containing native signal peptide. Transformed HEK-293 T cells were grown for 24 h in serum free medium, and the supernatant was then analysed by Western blotting for expression of histidine-tagged protein. The S1 protein was detected as a single 130 kDa band for both cells expressing S1 with native signal peptide and cells expressing S1 with TPA (Fig. 3a) . However, the presence of TPA substantially enhanced expression of S1. Therefore, S1 with TPA was subsequently used for production and purification of the vaccine antigen. Affinity purified S1 protein was analysed on SDS-PAGE by Coomassie blue staining and found to be of the expected size (Fig. 3b) . The yield of the S1 protein from HEK 293 T cell culture medium was found to be 30 mg/L, which was 10 and 100-fold higher than the yields obtained from insect cells or yeast cells respectively.