FIGURE 8.

Analysis of the secY Q212C mutant during OmpA-GFP-induced translocon jamming by disulfide cross-linking. A, view of T. maritima SecYEG docked in the back-to-back conformation with the same color scheme as described in the legend to Fig. 1. Magenta spheres indicate the Gln²¹² residue mutated to cysteine. B and C, the secY Q212C mutant was grown in the presence of 10 or 30 μm IPTG and 0.2% maltose until A₆₀₀ reached 0.15, when the OmpA-GFP chimera was induced by adding arabinose to a final concentration of 0.2%. Cells were harvested 45 min post jamming and treated with CuPhe₃ and DTT as indicated. B, Western blot of cell membranes probed with c-Myc antibody. C, Western blot of cells divided into cytoplasm/membrane (C) and periplasmic (P) fractions, compared with total (T) cell input, probed with MBP antibody.