Screening Of Goatpox Virus Receptors And Preliminary Study Of The Receptor Binding Proteins

Goatpox, caused by Goatpox virus (GTPV), is an acute and highly contagious disease, which isone of the OIE (World Organization for Animal Health) List diseases must be reported when occurred.GTPV infection leads to high morbidity and mortality causing huge economic losses to the sheepindustry, prompting increasing concern about research on GTPV.Currently, there is little information about the entry process and infection mechanism of GTPV.Virus entry into a host cell involves the specific interaction of virus with receptor molecules which areexpressed at the plasma membrane. The appropriate receptors existence on cell surface determine thecell susceptibility to viral infection. In the present study, phage display technology was used to screenthe high affinity and specific binding receptors for GTPV. The mRNA of primary cultured goattesticular cells was extracted and used as template for reverse transcription to get cDNA. Then thecDNA was connected with the phage vector and packaged in vitro to construct phage display library.After several rounds library screen with purified GTPV, the phage which can specifically bind to viruswas enriched and analyzed by sequencing. In this experiment, we successfully constructed the goattesticular cells cDNA phage display library, and the titer of the original packaging products is3.2×108pfu/mL, demonstrating that the library contains more than99%of the mRNA sequences. Twokinds of virus specific binding phage were obtained through screening of the library with purified virus.Sequence analysis revealed that the two gene fragments contained were homologous to sheep cysticfibrosis transmembrane regulatory protein and interleukin10receptor, respectively.Receptor binding protein is a kind of protein on the virus particle surface, which can specificallyrecognize and bind to virus receptors. In this study, bioinformatics methods were used to identify thespecific receptor binding proteins to GTPV receptors. The results showed that GTPV L1and L5proteinare homologous to the receptor binding proteins of vaccinia virus, with similarities of80%and70.2%for amino acid respectively. In order to clarify whether the GTPV L1and L5protein play a role in virusentry, inhibition assays were conducted using rabbit antiserum against the GTPV L1and L5proteinexpressed in E.coli cells. Results showed that anti-L1serum can effectively inhibit viruses infection.This study provides a method and basis for GTPV receptor research, and enriched the structuralproteins studies of GTPV, and laid the foundation for revealing of GTPV entry mechanism and targetedtherapy as well as prevention of goat pox.