Production of the recombinant hemagglutinin protein of the swine influenza virus A/H1N1 and analysis of its physicochemical and antigenic properties

Objectives. To analyze the physicochemical and antigenic properties of recombinant hemagglutinin protein of swine influenza virus strain A/H1N1 (swH1-His) obtained by transduction of suspension line HEK293 with recombinant human adenovirus serotype 5.

Methods. The de novo assembly of the target hemagglutinin gene was performed via the polymerase chain reaction. Recombinant adenovirus recAd5-swH1-His was obtained using the AdEasy™ Adenoviral Vector System kit. Accumulation of preparative amounts of recombinant protein was performed by transduction of recAd5-swH1-His suspension culture of HEK293 cells in a wave-type bioreactor. Recombinant hemagglutinin was isolated from the culture medium by metal-chelate affinity purification on a sorbent. The actual molecular mass and its correspondence to the expected value, as well as the presence of histidine residues were shown by electrophoresis and Western blot. The antigenic specificity of swH1-His was determined by indirect enzyme-linked immunosorbent assay with specific sera.

Results. Recombinant hemagglutinin swH1-His was obtained in the amount of 1.2 mg from 50 mL of culture fluid. The compliance of its mass with the declared molecular mass (≈70 kDa) was confirmed along with the presence of cross-linking with histidine residues. The antigenic specificity of swH1-His in reaction with sera was demonstrated.

Conclusions. The physicochemical and antigenic characteristics of recombinant protein hemagglutinin of swine influenza A/H1N1 (swH1-His) obtained by transduction of HEK293 cells with recombinant human adenovirus of serotype 5 were determined. The obtained recombinant hemagglutinin can be used as an antigen for animal and human influenza diagnostic purposes.

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