Figure 3.

NADPH‐linked peroxidase activity. (a) Determination of kinetic parameters (k _cat, K _M): reduction of H₂O₂ catalyzed by PRDX1 (black squares) or PRDX2 (red triangles) was monitored at different initial EcTrx1 concentrations with 200 μM NADPH, 1 μM EcTR, 0.5 μM PRDX1 or PRDX2 and 10 μM H₂O₂ in 50 mM phosphate buffer pH 7.4, 150 mM NaCl. NADPH oxidation was monitored at 340 nm. Experimental data was fitted to Michaelis–Menten equation (solid lines). (b) Representative time trace of H₂O₂‐dependent NADPH oxidation by the Prx/Trx/TR system at 50 μM H₂O₂. Reaction mixture contained 200 μM NADPH, 1 μM EcTR, 8 μM EcTrx1, 0.5 μM PRDX1 (black trace) or PRDX2 (red trace) in 50 mM phosphate buffer pH 7.4, 150 mM NaCl and the reaction was started by addition of H₂O₂. Dashed lines represent the initial slope of the reaction.