Expression of recombinant Influenza A H1N1 neuraminidase in Rachiplusia nu larvae

LAURA E. FALETTI; NICOLÁS URTASUN; ALEXANDRA MARISA TARGOVNIK; MARIANA B. ARREGUI; GUSTAVO J. LEVÍN; G. MARONICHE; FEDERICO JAVIER WOLMAN; OSVALDO CASCONE; MARÍA VICTORIA MIRANDA

Current Topics in Virology

Research Trends

Año: 2014 vol. 12 p. 65 - 75

0972-4591

Two recombinant baculoviruses carrying the fulllength or transmembrane-deleted neuraminidase (NA) genes of influenza were constructed with the aim to select the best strategy to express this viral antigenic protein in Rachiplusia nu larvae. These variants of NA were efficiently expressed with the expected molecular weight (∼60 kDa) in the Sf9 cell line. The transmembrane-deleted (Tm-less) NA variant was mainly detected in the supernatants and showed poor enzymatic activity due to the destabilization of the tetramer. The full-length NA variant integrity was confirmed by the presence of enzymatic activity and the yield in the supernatants was 6.57 ± 0.51 mg/l of culture. Although the full-length variant was not completely secreted, it happens to be the best option for NA expression. R. nu larvae produced 1.20 ± 0.20 mg of full-length recombinant NA/g larva. Thus, to produce 1 mg of NA, 153 ml of suspension culture with 1 x 106 cells/ml or only six larvae are needed. The full-length NA expressed in larvae was captured by a Concanavalin A affinity matrix, thus indicating the presence of glycosylated residues.