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Structural basis for TatA oligomerization: an NMR study of Escherichia coli TatA dimeric structure.

ABSTRACT: Many proteins are transported across lipid membranes by protein translocation systems in living cells. The twin-arginine transport (Tat) system identified in bacteria and plant chloroplasts is a unique system that transports proteins across membranes in their fully-folded states. Up to date, the detailed molecular mechanism of this process remains largely unclear. The Escherichia coli Tat system consists of three essential transmembrane proteins: TatA, TatB and TatC. Among them, TatB and TatC form a tight complex and function in substrate recognition. The major component TatA contains a single transmembrane helix followed by an amphipathic helix, and is suggested to form the translocation pore via self-oligomerization. Since the TatA oligomer has to accommodate substrate proteins of various sizes and shapes, the process of its assembly stands essential for understanding the translocation mechanism. A structure model of TatA oligomer was recently proposed based on NMR and EPR observations, revealing contacts between the transmembrane helices from adjacent subunits. Herein we report the construction and stabilization of a dimeric TatA, as well as the structure determination by solution NMR spectroscopy. In addition to more extensive inter-subunit contacts between the transmembrane helices, we were also able to observe interactions between neighbouring amphipathic helices. The side-by-side packing of the amphipathic helices extends the solvent-exposed hydrophilic surface of the protein, which might be favourable for interactions with substrate proteins. The dimeric TatA structure offers more detailed information of TatA oligomeric interface and provides new insights on Tat translocation mechanism.

SUBMITTER: Zhang Y

PROVIDER: S-EPMC4121141 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Structural basis for TatA oligomerization: an NMR study of Escherichia coli TatA dimeric structure.

Zhang Yi Y Hu Yunfei Y Li Hongwei H Jin Changwen C

PloS one 20140804 8

Many proteins are transported across lipid membranes by protein translocation systems in living cells. The twin-arginine transport (Tat) system identified in bacteria and plant chloroplasts is a unique system that transports proteins across membranes in their fully-folded states. Up to date, the detailed molecular mechanism of this process remains largely unclear. The Escherichia coli Tat system consists of three essential transmembrane proteins: TatA, TatB and TatC. Among them, TatB and TatC fo ...[more]

PMID: 25090434

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Project description:A lipopolysaccharide (LPS) molecule is a key component of the bacterial outer membrane used to protect the bacterium and to interact with the environment. To gain insight into its function, the study of the LPS conformation and dynamics at the molecular and cellular levels is necessary, but these highly diverse and dynamic membrane-LPS systems are difficult to study. In this work, by using NMR spectroscopy and molecular dynamics (MD) simulations, we determined the conformational preferences of an E. coli O176 O-antigen polysaccharide at the atomic level. Moreover, we analyzed the use of non-uniform sampling (NUS) for the acquisition of high dynamic range spectra, like 1H,1H-NOESY NMR experiments. A comparison of the effective transglycosidic distances derived from conventional uniformly sampled and NUS 1H,1H-NOESY data showed high similarity under equal measuring time conditions. Furthermore, the experimentally derived internuclear distances of the O-antigen polysaccharide with ten repeating units (RUs) showed very good agreement to those calculated from the MD simulations of the same O-antigen polysaccharide in solution. Analysis of the LPS bilayer simulations with five and with ten RUs revealed that, although similar with respect to populated states in solution, the O-antigen in LPS bilayers had more extended chains as a result of spatial limitations due to close packing. Additional MD simulations of O-antigen polysaccharides from E. coli O6 (branched repeating unit) and O91 (negatively charged linear repeating unit) in solution and LPS bilayers were performed and compared to those of O176 (linear polymer). For all three O-antigens, the ensemble of structures present for the polysaccharides in solution were consistent with the results from their 1H,1H-NOESY experiments. In addition, the similarities between the O-antigen on its own and as a constituent of the full LPS in bilayer environment makes it possible to realistically describe the LPS conformation and dynamics from the MD simulations.

Dataset Information

Structural basis for TatA oligomerization: an NMR study of Escherichia coli TatA dimeric structure.

Publications

Structural basis for TatA oligomerization: an NMR study of Escherichia coli TatA dimeric structure.

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