A screening assay for neuraminidase inhibitors using neuraminidases N1 and N3 from a baculovirus expression system

摘要

Context: Development of inexpensive and safe enzymatic assays to screen for putative neuraminidase inhibitors. Objective: Validate the use of recombinant neuraminidase expressed in baculovirus located on the viral surface capsule to develop a neuraminidase inhibitor screening assay. Materials and methods: Recombinant baculovirus particles displaying neuraminidase N1 and N3 were used as enzyme sources. The assay set-up required the use of 2′-(4-methylumbelliferyl)-α-D-acetyl neuraminic acid as substrate and oseltamivir carboxylate as benchmark inhibitor. Results: The assay was set up in a standard 96-well plate. The within- and between-assay coefficients of variation were, on average, less than 10%. The 50% inhibitory concentration values of the inhibitor were in good agreement with those determined by independent kinetic experiments. Discussion and conclusions: The assay showed satisfactory within- and between-assay repeatability. The obtained results suggest that recombinant baculovirus expressing neuraminidase located on the virus membrane capsule can be used to set up affordable and reliable neuraminidase inhibitors screening assays.