FIGURE 2.

Quantitation of the H₂O₂-oxidized cysteine residues of EF-G. A, quantitative analysis of thiol groups. Wild-type EF-G (2 μm) was treated with 5 mm DTT (reducing), 0.5 mm H₂O₂ (oxidizing), or the thiol-modifying (SH) reagent. The number of thiol groups per protein molecule of wild-type EF-G and its mutant derivatives was determined with DTNB. Values are mean ± S.D. (bars) of results from three independent experiments. B, the effect of H₂O₂ on the redox state of mutant derivatives of EF-G. After the derivatives of EF-G (2 μm) had been incubated with 0.5 mm H₂O₂ for 5 min, they were treated with the thiol-modifying reagent. Proteins were then fractionated by nonreducing SDS-PAGE. No, no modification with SH reagent; Red, reducing conditions; Ox, oxidizing conditions.