[Screening of polypeptides binding to porcine reproductive and respiratory syndrome virus by phage display library].

OBJECTIVE

To find specific polypeptides that bind porcine reproductive and respiratory syndrome virus (PRRSV) with high affinity and inhibit replication of PRRSV, we screened ligands on intact PRRSV virion by phage display library.

METHODS

The purified PRRSV was coated and then reacted with random peptide library displaying 12 amino acids fused on protein III of M13 phage. The selected peptides for target binding were assayed by ELISA after 3 rounds of biopanning and measured by 50% tissue culture infection dose. The positive clones with high affinity were used for automated sequencing, and the amino acid sequence of polypeptide displayed on phage was deduced. Synthesis of fluorescein isothiocyanate labelled polypeptide and establish a method for detection of PRRSV.

RESULTS

The enrichment was shown by ELISA after 3 rounds of biopanning and 17 positive clones bound to PRRSV with high affinity. Sequencing of the genes encoding these peptides in positive clones show some of conserved motifs. Two positive clones inhibited the replication of PRRSV in Marc-145 cells in vitro and decreased PRRSV TCID50 from 10(-7.3)/0.1 mL to 10(-3.2), 10(-3.6)/0.1 mL respectively. The fluorescein isothiocyanate labelled peptide was able to detect PRRSV at the concentration of 5 mg/L.

CONCLUSION

Positive clones against PRRSV can be selected from phage display peptide library and so provide a potential tool for highly sensitive diagnostic kits and novel antiviral agents.