FIGURE 9.

Sulfenylation and disulfide bond formation kinetics of HSPB1 in response to peroxisomal H₂O₂ production. Flp-In T-REx 293 cells expressing po-DD-DAO and containing c-IBD-SBP-YAP1C (A) or not (B,C) were transfected (B,C) or not (A) with a plasmid encoding no EGFP-fusion protein (−), EGFP-PEX11B, or EGFP-HSPB1. The cells were incubated in DPBS containing 10 mM 3-AT and 10 mM D-Ala. At the indicated time points, the cells were processed as detailed in the legend of (A) Figure 5, or (B,C) processed for SDS-PAGE under non-reducing (-β-ME) or reducing (+β-ME) conditions and subsequently subjected to immunoblot analysis with antibodies specific for EGFP. The migration points of relevant molecular mass markers (expressed in kDa) are shown on the left. The arrows and arrowheads mark the non-modified and oxidatively modified proteins, respectively. Note that panel B was included to document the non-specific immunoreactive bands (marked by asterisks) of the anti-EGFP antiserum.