MIRANDA Maria Victoria
Expression of recombinant influenza A H1N1 neuraminidase in Rachiplusia nu larvae
LAURA FALETTI; URTASUN NICOLÁS; ALEXANDRA TARGOVNIK; MARIANA BERNADETT ARREGUI; GUSTAVO LEVIN; MARONICHE G.; WOLMAN FEDERICO; CASCONE OSVALDO; MARÍA VICTORIA MIRANDA
Current topics in Virology
Año: 2014 vol. 12 p. 65 - 65
Two recombinant baculoviruses carrying the full-length 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 culture. Although the full-length variant was not completely secreted, it becomes 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 1x106 cell/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.