Figure 2.
Formation of a common intermediate during GSH-dependent detglutathionylation of ceCblC-GSCbl or dealkylation of ceCblC-MeCbl. An intermediate with a λmax = 507 nm, assigned as [ceCblC-GSCbl-GS−], is observed in A–C. A, changes in the absorption spectrum of ceCblC (50 μm)-bound GSCbl (30 μm) mixed with 4 mm GSH under anaerobic conditions. Black trace, 0 s (λmax = 413 nm); red trace, 0.2 s (λmax = 507 nm); blue trace, 2.0 s (λmax = 389 nm); gray traces represent intermediate time points. The inset shows the time-dependent changes in absorbance at 389 nm (red) and 507 nm (black). B, spectral changes upon mixing ceCblC (50 μm)-bound GSCbl (30 μm) with 4 mm GSH under aerobic conditions. Black trace, 0 s (λmax = 413 nm); red trace, 0.2 s (λmax = 501–510 nm); blue trace, 7.0 s (λmax = 475 nm); orange trace, 20 s (λmax = 389 nm); gray traces represent intermediate time points. The inset shows the time-dependent changes in absorbance at 389 nm (red), 473 nm (blue), and 507 nm (black). C, change in the absorption spectra of ceCblC (50 μm)-bound MeCbl (30 μm) incubated with 4 mm GSH under aerobic conditions. Black trace, 0 s (λmax = 454 nm); red trace, 4.0 s (λmax = 487 nm); blue trace, 7.5 s (λmax = 479 nm); orange trace, 20 s (λmax = 389 nm); gray trace, intermediate time points. Inset, time-dependent changes in absorbance at 389 nm (red), 473 nm (blue), and 507 nm (black).