Expression and immunogenicity of secreted forms of bovine ephemeral fever virus glycoproteins applied to subunit vaccine development

Aims: Vaccines for bovine ephemeral fever virus (BEFV) are available but are difficult to produce, expensive or suffer from genetic instability. Therefore, we designed constructs encoding C-terminally truncated forms (transmembrane anchoring region deleted) of glycoproteins G and G_NS such that they were secreted from the cell into the media to achieve high-level antigen expression, correct glycosylation pattern and enable further simple purification with the V5 epitope tag.

Methods and results: In this study, synthetic biology was employed to create membrane-bound and secreted forms of G and G_NS glycoprotein. Mammalian cell culture was employed as an antigen expression platform, and the secreted forms of G and G_NS protein were easily purified from media using a highly effective, single-step method. The V5 epitope tag was genetically fused to the C-termini of the proteins, enabling detection of the antigen through immunoblotting and immunomicroscopy. Our data demonstrated that the C-terminally truncated form of the G glycoprotein was efficiently secreted from cells into the cell media. Moreover the immunogenicity was confirmed in mice test.

Conclusions: The immuno-dot blots showed that the truncated G glycoprotein was present in the total cell extract, and was clearly secreted into the media, consistent with the western blotting data and live-cell images. Our strategy presented the expression of secreted, epitope-tagged, forms of the BEFV glycoproteins such that appropriately glycosylated forms of BEFV G protein was secreted from the BHK-21 cells. This indicates that high-level expression of secreted G glycoprotein is a feasible strategy for large-scale production of vaccines and improving vaccine efficacy.

Significance and impact of the study: The antigen expression strategy designed in this study can produce high-quality recombinant protein and reduce the amount of antigen used in the vaccine.