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. 2011 Nov 2;6(11):e26974. doi: 10.1371/journal.pone.0026974

Figure 3. NADPH-linked reductase activity assay of recombinant MsrA and MsrB from S. Typhimurium.

Figure 3

A. The graph shows the amount of oxidized NADPH per minute for the reduction of 500, 100, 50 and 5 µM methyl p-tolyl sulfoxide by MsrA and MsrB, respectively. B. The graph shows the amount of oxidized NADPH per minute for the reduction 500, 125 and 50 µM N-acetyl methionine sulfoxide by MsrA and MsrB, respectively. The decrease of absorbance at 340 nm due to NADPH oxidation was measured in the presence of the S- or R-isomer of methyl p-tolyl sulfoxide (A), and in the presence of the S- or R-isomere of N-acetyl methionine sulfoxide (B) using purified His-tagged MsrA or MsrB. Besides 10 µM purified MsrA or MsrB, the test system contained 10 µM TrxR, 10 µM TrxB, 450 µM NADPH, and various concentrations of S- or R-methyl p-tolyl sulfoxide (Sigma), and N-acetyl methionine sulfoxide (MOLISA), respectively. Reactions were filled up to a final volume of 500 µl with a buffer containing 50 mM HEPES and 1 mM EDTA (pH 7.4). Reactions were carried out at 25°C and started after 10 min pre-incubation by addition of the oxidized methionine derivatives.