Abstract
Twin-arginine translocation (Tat) systems mediate the transmembrane translocation of completely folded proteins that possess a conserved twin-arginine (RR) motif in their signal sequences. Many Tat systems consist of three essential membrane components named TatA, TatB, and TatC. It is not understood why some bacteria, in addition, constitutively express a functional paralog of TatA called TatE. Here we show, in live Escherichia coli cells, that, upon expression of a Tat substrate protein, fluorescently labeled TatE-GFP relocates from a rather uniform distribution in the plasma membrane into a number of discrete clusters. Clustering strictly required an intact RR signal peptide and the presence of the TatABC subunits, suggesting that TatE-GFP associates with functional Tat translocases. In support of this notion, site-specific photo cross-linking revealed interactions of TatE with TatA, TatB, and TatC. The same approach also disclosed a pronounced tendency of TatE and TatA to hetero-oligomerize. Under in vitro conditions, we found that TatE replaces TatA inefficiently. Our collective results are consistent with TatE being a regular constituent of the Tat translocase in E. coli.
Highlights
Twin-arginine translocases, which transport folded proteins across cellular membranes, often consist of three proteins, TatA, TatB, and TatC, but sometimes have an additional tatE gene alone (TatE) available
Our collective results are consistent with TatE being a regular constituent of the Tat translocase in E. coli
To learn more about the role TatE might play in Tat-dependent protein translocation, we undertook a combined in vivo and in vitro analysis of how TatE associates with the Tat translocase of E. coli
Summary
Twin-arginine translocases, which transport folded proteins across cellular membranes, often consist of three proteins, TatA, TatB, and TatC, but sometimes have an additional TatE available. Many Tat systems consist of three essential membrane components named TatA, TatB, and TatC It is not understood why some bacteria, in addition, constitutively express a functional paralog of TatA called TatE. Clustering strictly required an intact RR signal peptide and the presence of the TatABC subunits, suggesting that TatE-GFP associates with functional Tat translocases In support of this notion, site-specific photo cross-linking revealed interactions of TatE with TatA, TatB, and TatC. One hypothesis suggests that TatA associates with the TatBC receptor complex, undergoes reorganization, and oligomerizes to form a translocation channel for the substrate (summarized in Ref. 4). To learn more about the role TatE might play in Tat-dependent protein translocation, we undertook a combined in vivo and in vitro analysis of how TatE associates with the Tat translocase of E. coli
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