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Expression and immunogenicity of secreted forms of bovine ephemeral fever virus glycoproteins applied to subunit vaccine development
Ist Teil von
Journal of applied microbiology, 2021-09, Vol.131 (3), p.1123-1135
Ort / Verlag
England: Oxford University Press
Erscheinungsjahr
2021
Quelle
Wiley-Blackwell Journals
Beschreibungen/Notizen
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 GNS 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 GNS glycoprotein. Mammalian cell culture was employed as an antigen expression platform, and the secreted forms of G and GNS 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.