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. 2015 Sep 2;5:13666. doi: 10.1038/srep13666

Figure 5. Inhibitory activity.

Figure 5

(a,b) SDS-Page of 50 μM WT NS and 50 μM E289A NS incubated with 22 μM tPA at 23 °C for different time intervals (as reported in the figure); the first lanes are the Molecular Weight markers (M), the second lanes are the native NS alone (NS). An arrow indicates the different species: double chain (dc) tPA/NS Complex, single chain (sc) tPA/NS Complex, single chain (sc) tPA, native NS, cleaved NS. The dc-Complex originates from the residual dc-tPA, and it looses the not bonded chain upon denaturation in reducing conditions. Analogously, the two chain of dc-tPA are separated upon denaturation with SDS and run with different velocity in the gel lanes (below 37 kDa). (c) Progress curves of hydrolysis of IPR-pNA (0.2 mM) by tPA (1 nM) in the presence of 0 NS (black line), 15 nM E289A NS (dotted violet line), 15 nM WT NS (solid violet line), 45 nM E289A NS (dotted blue line), 45 nM WT NS (solid blue line). (d) Band densities from SDS PAGE as in panel (a) for WT NS (solid lines) and panel (b) for E289A NS (dashed lines): NS/tPA Complex (blue lines), cleaved NS (green lines), native NS (brown lines). Points are normalised by the total density value of each lane. The points are averages over triplicate experiments.