The Expression Of Recombinant GP5 Proteins Deleted Transmembrane Domains And The Study Of Their Immune Effect

The envelope glycoprotein 5 (GP5) is one of the major structural proteins of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV). In this report, expression of GP5 has been improved by deleting the very hydrophobic transmembrane regions of envelope glycoprotein gene (ORF5) and A epitope. The ORF5 gene segment with the removing of N-terminal transmembrane region and the other ORF5 gene segment deleting the N-terminal and middle transmembrane regions at the same time, were cloned into Prokaryotic expression vector pET-32 a(+), named pET-ORF5-1 and pET-ORF5-2 respectively. The expressing level of the deletion mutant ORF5-2 was about 3 times higher than the deletion mutant ORF5-1. The two recombinant fusion proteins were recognized by Western-blot with pig positive sera. This studies lay foundations for further study on the biological founction of GP5 protein and the development of vaccines for PRRSV.To study the immunological properties of fused expression products of PRRSV orf5 gene segments in pET32a(+) vector, and lay the foundation for developing PRRSV genetic engineering sub unit vaccines. The recombinant GP5-1 and GP5-2 proteins which were deleted transmembrane domains and exposed B epitope, were expressed. Mice were injected with the proteins. The humoral and cellular immunological effects were identified by ELISA, micro cell culture neutralization test and lymphocytes stimulation test. In immunized mice, specific antibodies with the highest titre of 1:1600 were observed, which could specifically bind to GP5-land GP5-2 proteins. No detectable neutralizing antibodies were tested in all of the immunized mice. The level of spleen lymphocyte proliferation was measured by MTT. The results indicated products of orf5 gene segments are immunogenic, which can stimultaneously stimulate humoral and cellular immunological responses against GP5-1 and GP5-2 proteins of PRRSV.