Efficient twin arginine translocation (Tat) pathway transport of a precursor protein covalently anchored to its initial cpTatC binding site

The thylakoid twin arginine protein translocation ( Tat) system operates by a cyclical mechanism in which precursors bind to a cpTatC-Hcf106 receptor complex, which then recruits Tha4 to form the translocase. After translocation, the translocase disassembles. Here, we fine-mapped initial interactions between precursors and the components of the receptor complex. Precursors with (Tmd) Phe substitutions in the signal peptide and early mature domain were bound to thylakoids and photo-cross-linked to components. cpTatC and Hcf106 were found to interact with different regions of the signal peptide. cpTatC cross-linked strongly to residues in the immediate vicinity of the twin arginine motif. Hcf106 cross-linked less strongly to residues in the hydrophobic core and the early mature domain. To determine whether precursors must leave their initial sites of interaction during translocation, crosslinked precursors were subjected to protein transport conditions. tOE17 cross-linked to cpTatC was efficiently translocated, indicating that the mature domain of the precursor can be translocated while the signal peptide remains anchored to the receptor complex.