Twin-arginine translocation-arresting protein regions contact TatA and TatB.

Tat systems translocate folded proteins across biological membranes of prokaryotes and plant plastids. TatBC complexes recognize N-terminal Tat signal peptides that contain a sequence motif with two conserved arginines (RR-motif), and transport takes place after a recruitment of TatA. Unfolded Tat substrate domains lower translocation efficiency and too long linkers lead to translocation arrest. To identify the components that interact with transported proteins during their passage through the translocon, we used a Tat substrate that arrests translocation at a long unfolded linker region, and we chose in vivo site-directed photo cross-linking to specifically detect the interactions of this linker region. For comparison, we included the interactions of the signal peptide and of the folded domain at the C-terminus of this construct. The data show that the linker contacts only two, structurally similar Tat components, namely TatA and TatB. These contacts depend on the recognition of the Tat-specific signal peptide. Only when membrane translocation of the globular domain was allowed--i.e., in the absence of the linker--we observed the same TatAB-contacts also to the globular domain. The data thus suggest that mature protein domains are translocated through a TatAB environment.