Fig. 3

Analysis of S-nitrosation on PfGluPho (a), PfG6PD (b), and PvG6PD (c). Pre-reduced enzymes were incubated with 0 or 1 mM GSNO for 1 h at 22 °C in the dark and subjected to the biotin switch assay. Samples without sodium ascorbate served as a control. S-nitrosylated proteins were detected by anti-biotin western blot. Coomassie staining of the samples used for western blotting served as a loading control. Representative blots from at least two independent experiments are shown. To study the impact of S-nitrosation on enzyme activity, pre-reduced enzymes were incubated with different concentrations of GSNO (0–1000 µM) for 180 min at 22 °C in the dark. Activity of each sample was measured directly after adding GSNO, as well as after 30, 60, 90, 120, and 180 min. Activity of the sample with 0 µM GSNO was defined as 100%. Values shown are mean ± SD from at least three independent determinations using two different batches of enzyme