Screening And Identification Of Affinity Peptide To Vp2Protein Of Procine Parvovirus

Porcine parvovirus (PPV) causes reproductive failure in swine, The virus is ubiquitous among swine throughout the world and is enzootic in most herds that have been tested.The VP2protein of PPV is a major structural protein of virus and has specific immunogenicity. The VP2gene is a conserved sequence in the viral genome.The expressed product would be used in diagnosis.According to the VP2gene sequence of PPVstrain BQ published in GenBank, a pair of primers was used to amplify the VP2gene by PCR. The VP2consists of1740bp in length. VP2was digested BamH I and SalI then inserted into the downstream of T7promoter of prokaryotic expression vector pET32a(+) to construct an expression plasmid VP2-pET32a. After induction by IPTG, the fusion protein was highly expressed in Escherichia Coli Rosseta (DE3)in the form of inclusion bodies. SDS-PAGE indicates that the VP2protein is approximately82KDa as expected. Western blot indicates that the protein can be recognized by swine anti-PPV serum. All results indicated that the recombinant fusion proteincan be used as an antigen of diagnostic assay to detect the PPV antibody in serum.Phage display is a technology for ligand identification to target protein. In this study, Random12-mer Phage Display Peptide Library was used to panning the VP2protein for five rounds. In the four round, tagged protein of pET32a(+) was used for a subtraction panning.Finally, ten monoclonal phages were selected randomly and subjected to sequence comparison. The results showed that six different peptides were screened by panning on the VP2recombinat protein. Two peptides that have the highest binding activity to the VP2were synthesized. Their consensus sequences are HTHQLHFLNHNP named1and HNLHLYHYLRSA named2, respectively. The inhibition of the two phages bearing either1or2peptide to PPV were evaluated in ST cell by MTT and Immunofluorescence.The results indicated PPV and peptide incubation led to a decreased infectivity of PPV. The ability of neutralization of peptide2was better than peptide1, and combination of peptides1+2gave rise to the most potent inhibition to PPV. The current study may be helpful for design preventive strategies to PPV infection based on phage display techniques.