Transient expression of a bovine leukemia virus envelope glycoprotein in plants by a recombinant TBSV vector

Plants offer a unique combination of advantages for the production of valuable recombinant proteins in a relatively short time. For instance, a variety of diagnostic tests have been developed that use recombinant antigens expressed in plants. The envelope glycoprotein gp51 encoded by Bovine leukemia virus (BLV) is one of the essential subunits for viral infectivity. It was indicated that the recombinant gp51 (rgp51) of BLV сan be used as an synthetic alternative antigen useful in the diagnosis of BLV infection in cattle. Here we evaluate the potential for using a viral vector based on the genome of Tomato bushy stunt virus (TBSV) for the efficient expression of BLV envelope glycoprotein rgp51 in Nicotiana benthamiana plants. The codon-optimized gene encoding rgp51 was synthesized by the de novo DNA synthesis to replace the GFP gene in the TBSV-derived viral vector that was then delivered into 4-5 week old N. benthamiana plants by agroinfiltration. Expression of recombinant his-tagged rgp51 was verified by protein extraction followed by western blot procedures, and by purification using Ni²⁺-affinity chromatography. The molecular weight of this plant-expressed rgp51 ranged from 43 to 55 kDa and it was shown to be glycosylated. Important for potential use in diagnostic tests, purified rgp51 specifically reacted with BLV infected bovine sera while no reaction was observed with the negative serum samples.