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Figure 2

From: Adaptive mutations of neuraminidase stalk truncation and deglycosylation confer enhanced pathogenicity of influenza A viruses

Figure 2

In vitro evaluation of the rK/09 and its NA stalk mutant viruses. (A) rK/09 and its NA stalk mutant viruses (rΔ53–60, rNN58SS, and rN58T) were generated by reverse genetics based on the NA stalk truncation or deglycosylation strategy. (B) In the western blotting assay, NA stalk mutations (truncation and deglycosylation) were confirmed based on the mobility shift of NA proteins. (C) The enzyme kinetics data were fit to the Michaelis-Menten equation by nonlinear regression to determine the Michaelis constant (K m ) and maximum velocity (V max ) of substrate conversion. Results were given as the means ± SD from three independent determinations on triplicate samples where the R 2 was > 0.99. (D) Replication properties of each virus were evaluated in MDCK and A549 cells at a multiplicity of infection (MOI) of 0.001. The results were represented as the mean values from three independent experiments. Error bars denote SD. * P < 0.05, ** P < 0.01, and *** P < 0.001 for the results assessed in the MDCK cells and ≠ P < 0.05 and ≠≠≠ P < 0.001 in the A549 cells (compared with the rK/09 replication titers).

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