FIGURE 3.

Cross-linking of RR precursors to TatC requires an intact RR motif and occurs independently of the proton-motive force. Experimental details are as in the legend for Fig. 2. A, cross-linking to four prominent contact sites of TatC is compared between wild-type TorA-mCherry (RR) and a mutant derivative, in which the RR pair had been exchanged against two lysines (KK). Tat⁺-INV were included as Bpa-free control vesicles. B, in vitro synthesis of TorA-MalE in the presence of the indicated INV. Where indicated, CCCP was added to dissipate the proton-motive force of the INV. Control samples received an equal amount of the solvent Me₂SO. The addition of CCCP resulted in a block in translocation, i.e. production of the mature form (m) of TorA-MalE but did not influence cross-linking to TatC (stars). C, as in B except for using TorA-mCherry as RR precursor. For unknown reasons, the TatC adducts to various precursors often appeared as double bands (B and C). The size difference is too small to result from cross-linking to precursors and the signal sequence-less forms.