Fig 5. Effects of TrxM on the redox state of RpaA, RpaB and ManR.

Purified TFs were treated with oxidizing or reducing agents, modified with NEM or PEG-maleimide, fractionated by non-reducing 15% (for NEM) or 12% (for PEG maleimide) SDS-PAGE and stained with CBB. RpaA modified with NEM (A) or PEG-maleimide (B), RpaB modified with NEM (C) or PEG-maleimide (D), ManR modified with NEM (E) or PEG-maleimide (F). TFs (5 μM) were incubated with 100 mM DTT, H₂O₂ (1 or 10 mM) or aldrithiol (100 μM). Oxidized TFs were reduced by 100 mM DTT or TrxM (0.5 or 5 μM) with indicated concentration of DTT. After precipitation with 10% (w/v) trichloroacetic acid, TFs were subjected to the thiol modification using NEM or PEG-maleimide. Ox and Red indicate the oxidized and reduced forms of a TF, respectively.