Screening And Characterization Of S1 Protein Affinity Peptides Of Porcine Epidemic Diarrhea Virus

Porcine epidemic diarrhea (PED) is a highly contagious, enteric disease of swine caused by porcine epidemic diarrhea virus (PEDV). Since 1978, the outbreak of PED in Europe and many Asian countries have led to huge economic losses in pig industry. Thus, there is a urgent need to understand its pathological mechanism and to develop novel PED vaccine for the prevention of PED.The S protein of PEDV is a glycoprotein localized on the virion surface, which plays a key role in the specific receptor binding, cell membrane fusion, and induction of neutralizing antibody for its possessing neutralizing epitopes and receptor binding domain. Given this, screening and identification of receptor binding domain and antigenic epitopes were carried out in this study. It will provide valuable information for development of diagnosis technology, novel vaccine, and designing antiviral immunization strategy of PED.The experiment successfully constructed a recombinant expression vector pET-30a-S1. High level expression of S1 protein was analyzed by SDS-PAGE at different time at 37℃after IPTG induction. The S1 protein was purified and injected to a rabbit to generate polyclonal antibody. Western-blotting result showed polyclonal antibody could specifically bind to PEDV S1 protein, which indicated that recombinant fusion protein has excellent immunogenicity.Random 12-mer Phage Display Peptide Library was used to panning the S1 protein for four rounds. Ten monoclonal phages were selected randomly and subjected to sequence comparison. The results showed that three different peptides were screened by panning on the S1 recombinat protein. Three peptides that have the highest binding activity to S1 were synthesized. Their sequences are MPAVMSSAQVPR named M, NLSNRLNLSPGI named N and YVIHQPYAMALR named Y, respectively. ELISA showed that both peptides could bind to S1 protein and inhibit cell infection by PEDV in vitro. The immunized Kunming mice were used to detect the anti-viral activity of peptides in vivo. The results showed that the peptide M and N could neutralize viral infection activity. The current study should be helpful for establishing specific diagnosis method and developing new subunit vaccine for PEDV.