Abstract
The twin-arginine translocation (Tat) system is an integral membrane protein complex that accomplishes the remarkable feat of transporting large, fully folded polypeptides across the inner membrane of bacteria, into the periplasm. In Escherichia coli, Tat comprises three membrane proteins: TatA, TatB and TatC. How these proteins arrange themselves in the inner membrane to permit passage of Tat substrates, whilst maintaining membrane integrity, is still poorly understood. TatA is the most abundant component of this complex and facilitates assembly of the transport mechanism. We have utilised immunogold labelling in combination with array tomography to gain insight into the localisation and distribution of the TatA protein in E. coli cells. We show that TatA exhibits a uniform distribution throughout the inner membrane of E. coli and that altering the expression of TatBC shows a previously uncharacterised distribution of TatA in the inner membrane. Array tomography was used to provide our first insight into this altered distribution of TatA in three-dimensional space, revealing that this protein forms linear clusters in the inner membrane of E. coli upon increased expression of TatBC. This is the first indication that TatA organisation in the inner membrane alters in response to changes in Tat subunit stoichiometry.
Highlights
The bacterial twin-arginine translocation (Tat) system transports fully folded proteins across the bacterial inner membrane and the chloroplast thylakoid membrane to the periplasm
We have developed a novel combination of immunogold labelling and array tomography [28] of E. coli to unveil the distribution of TatA in the inner membrane (Supplementary Figure S1)
Tata exhibits a uniform distribution in the inner membrane of E. coli in cells overexpressing only TatA
Summary
The bacterial twin-arginine translocation (Tat) system transports fully folded proteins across the bacterial inner membrane and the chloroplast thylakoid membrane to the periplasm (reviewed by [1,2]). In Gram-negative bacteria, the Tat system comprises three membrane proteins: TatA, TatB and TatC [5,6]. Substrates are targeted to the Tat machinery via an N-terminal signal peptide, which contains an invariant twin-arginine motif [7,8]. At the target membrane the substrate makes contact with a TatBC complex containing multiple copies of these subunits [9,10]. Interaction with this substrate-binding complex triggers the recruitment of a separate TatA complex [11,12] to form the active translocase through which the substrate can pass. The precise mechanism of translocation is, poorly understood and the full translocation system has not been characterised
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