Figure 3.

Interaction between the oxidized form of the PrxQ and Trxh1_CS proteins. (A) Same experiment as presented in Figure 2A, except using PrxQ as the analyte. (B) Trxh1_CS (50μM) and PrxQ (50μM) were incubated for the periods indicated. The reaction was then quenched with 5% TCA (w/v). Proteins were separated by non-reducing SDS-PAGE. (C) Band intensities of the mixed disulfide intermediate complexes (intermediate), 45 kDa complexes, and the Trxh1_CS dimers (Trx dimer) obtained from SDS-PAGE presented in panel (B) were determined using a Scion image software and plotted. a.u.; arbitrary units. (D) Trxh1_CS (50μM) and oxidized PrxQ (50μM) were incubated for 3 min at room temperature, and the complexes that formed were then incubated with 10 mM NEM. The protein mixture was then loaded onto G2000SW_XL and separated. (E) Same experiment as in (D) except for incubation in the absence of NEM.