FIGURE 3.

Overoxidation of purified Anabaena 2-Cys Prx in vitro. A, purified recombinant wild type and mutant versions of Anabaena 2-Cys Prx (4 μm final concentration) was incubated for 5 min in the absence or presence of 0.5 mm hydrogen peroxide. Thereafter, DTT was added to each sample at a final concentration of 10 mm, and samples were incubated for another 10 min and precipitated with 5% trichloroacetic acid. Samples (1 μg of protein) were subjected to two-dimensional gel electrophoresis and proteins visualized with Coomassie Brilliant Blue. Cont, control; red, reduced; ox, overoxidized. B and C, MALDI-TOF analysis of mass-to-charge ratios (m/z) of mutant versions, as indicated, of 2-Cys Prx from Anabaena. Continuous lines, control; dashed lines, H₂O₂-treated. Intens. [a.u.], intensity (arbitrary units).