Abstract

The goose parvovirus (GPV) Rep1 protein is both essential for viral replication and a potential target for GPV diagnosis, but its protein characterization and intracellular localization is not clear. We constructed a recombinant plasmid, pET28a/GPV-Rep1, and expressed the Rep1 gene in BL21 (DE3) Escherichia coli. A protein approximately 75 kDa in size was obtained from lysates of E. coli cells expressing the recombinant plasmid. SDS-PAGE analysis showed that after induction with 0.6 mM isopropyl β-D-thiogalactosidase (IPTG) at 30°C for 5 h, the Rep1 protein was highly overexpressed. Two methods used to purify proteins, a salinity-gradient elution and Ni-NTA affinity chromatography, were performed. The amount of Rep1 protein obtained by Ni-NTA affinity chromatography was 41.23 mg, while 119.9 mg of Rep1 protein was obtained by a salinity-gradient elution from a 1 L E. coli BL21 (DE3) culture. An immunogenicity analysis showed that the protein could significantly elicit a specific antibody response in immunized goslings compared to control groups. Antibody titers peaked to 1:5120 (optical density (OD) 450 = 3.9) on day 28 after immunization but had mean titers of 1:10,240 (OD450 = 4.2) in gosling groups immunized with a commercially available GPV-attenuated vaccine strain. Experiments examining subcellular localization showed that the Rep1 protein appeared to associate predominantly with the nuclear membrane, especially during later times of infection. This work provides a basis for biochemical and structural studies on the GPV Rep1 protein.

Keywords: Goose parvovirus, Rep1 protein, salinity-gradient elution, intracellular localization.