Abstract
The twin-arginine protein translocation system (Tat) transports folded proteins across the bacterial cytoplasmic membrane and the thylakoid membranes of plant chloroplasts. The Tat transporter is assembled from multiple copies of the membrane proteins TatA, TatB, and TatC. We combine sequence co-evolution analysis, molecular simulations, and experimentation to define the interactions between the Tat proteins of Escherichia coli at molecular-level resolution. In the TatBC receptor complex the transmembrane helix of each TatB molecule is sandwiched between two TatC molecules, with one of the inter-subunit interfaces incorporating a functionally important cluster of interacting polar residues. Unexpectedly, we find that TatA also associates with TatC at the polar cluster site. Our data provide a structural model for assembly of the active Tat translocase in which substrate binding triggers replacement of TatB by TatA at the polar cluster site. Our work demonstrates the power of co-evolution analysis to predict protein interfaces in multi-subunit complexes.
Highlights
Protein export across the cell membrane of prokaryotes occurs through two parallel pathways
It is notable that the unusually short TatA transmembrane helix (TMH) is well-matched in length to the section of the kinked TatC TM5 with which it primarily interacts
Our structural model suggests that the polar cluster residues mediate complex formation between TatA/TatB and TatC. To test this idea we investigated the effect of polar cluster defects on proteinprotein interactions within the E. coli translocation system (Tat) system
Summary
Protein export across the cell membrane of prokaryotes occurs through two parallel pathways. In prokaryotes the requirement for a functional Tat pathway varies with the organism and their growth environment (Palmer and Berks, 2012). Even under permissive growth conditions, loss of the Tat pathway results in serious pleiotropic effects on major cellular processes including energy metabolism, nutrient acquisition, virulence, and formation of the cell envelope (Berks et al, 2003; De Buck et al, 2008; Palmer and Berks, 2012). The Tat transport system has been evolutionarily conserved in plant chloroplasts where it mediates protein import across the thylakoid membrane and is essential for the formation of a functional photosynthetic apparatus (Celedon and Cline, 2013)
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