Structure of the TatC core of the twin-arginine protein transport system


The twin-arginine translocation (Tat) pathway is one of two general protein transport systems found in the prokaryotic cytoplasmic membrane and is conserved in the thylakoid membrane of plant chloroplasts. The defining, and highly unusual, property of the Tat pathway is that it transports folded proteins, a task that must be achieved without allowing appreciable ion leakage across the membrane. The integral membrane TatC protein is the central component of the Tat pathway. TatC captures substrate proteins by binding their signal peptides. TatC then recruits TatA family proteins to form the active translocation complex. Here we report the crystal structure of TatC from the hyperthermophilic bacterium Aquifex aeolicus. This structure provides a molecular description of the core of the Tat translocation system and a framework for understanding the unique Tat transport mechanism.

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Figure 1: Purified A. aeolicus TatC binds Tat signal peptides.
Figure 2: Structure of A. aeolicus TatC.
Figure 3: Identification of the signal peptide binding site on TatC.
Figure 4: Sites of interaction with other Tat components.
Figure 5: A conceptual model for the environment of TatC in the substrate-bound translocation site.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

The coordinates and experimental data have been deposited at the Protein Data Bank with accession code 4b4a.


  1. 1

    Park, E. & Rapoport, T. A. Mechanisms of Sec61/SecY-mediated protein translocation across membranes. Annu. Rev. Biophys. 41, 21–40 (2012)

    CAS  Article  Google Scholar 

  2. 2

    Palmer, T. & Berks, B. C. The twin-arginine translocation (Tat) protein export pathway. Nature Rev. Microbiol. 10, 483–496 (2012)

    CAS  Article  Google Scholar 

  3. 3

    Frobel, J., Rose, P. & Muller, M. Twin-arginine-dependent translocation of folded proteins. Phil. Trans. R. Soc. Lond. B 367, 1029–1046 (2012)

    Article  Google Scholar 

  4. 4

    Celedon, J. M. & Cline, K. Intra-plastid protein trafficking; how plant cells adapted prokaryotic mechanisms to the eukaryotic condition. Biochim. Biophys. Acta (2012)

  5. 5

    Berks, B. C., Palmer, T. & Sargent, F. The Tat protein translocation pathway and its role in microbial physiology. Adv. Microb. Physiol. 47, 187–254 (2003)

    CAS  Article  Google Scholar 

  6. 6

    De Buck, E., Lammertyn, E. & Anne, J. The importance of the twin-arginine translocation pathway for bacterial virulence. Trends Microbiol. 16, 442–453 (2008)

    CAS  Article  Google Scholar 

  7. 7

    Barkan, A., Miles, D. & Taylor, W. C. Chloroplast gene expression in nuclear, photosynthetic mutants of maize. EMBO J. 5, 1421–1427 (1986)

    CAS  Article  Google Scholar 

  8. 8

    Settles, A. M. et al. Sec-independent protein translocation by the maize Hcf106 protein. Science 278, 1467–1470 (1997)

    ADS  CAS  Article  Google Scholar 

  9. 9

    Sargent, F. et al. Overlapping functions of components of a bacterial Sec-independent protein export pathway. EMBO J. 17, 3640–3650 (1998)

    CAS  Article  Google Scholar 

  10. 10

    Bogsch, E. G. et al. An essential component of a novel bacterial protein export system with homologues in plastids and mitochondria. J. Biol. Chem. 273, 18003–18006 (1998)

    CAS  Article  Google Scholar 

  11. 11

    Hu, Y., Zhao, E., Li, H., Xia, B. & Jin, C. Solution NMR structure of the TatA component of the twin-arginine protein transport system from gram-positive bacterium Bacillus subtilis. J. Am. Chem. Soc. 132, 15942–15944 (2010)

    CAS  Article  Google Scholar 

  12. 12

    Berks, B. C. A common export pathway for proteins binding complex redox cofactors? Mol. Microbiol. 22, 393–404 (1996)

    CAS  Article  Google Scholar 

  13. 13

    Chaddock, A. M. et al. A new-type of signal peptide—central role of a twin-arginine motif in transfer signals for the ΔpH-dependent thylakoidal protein translocase. EMBO J. 14, 2715–2722 (1995)

    CAS  Article  Google Scholar 

  14. 14

    Stanley, N. R., Palmer, T. & Berks, B. C. The twin arginine consensus motif of Tat signal peptides is involved in Sec-independent protein targeting in Escherichia coli. J. Biol. Chem. 275, 11591–11596 (2000)

    CAS  Article  Google Scholar 

  15. 15

    Alami, M. et al. Differential interactions between a twin-arginine signal peptide and its translocase in Escherichia coli. Mol. Cell 12, 937–946 (2003)

    CAS  Article  Google Scholar 

  16. 16

    Gerard, F. & Cline, K. Efficient twin arginine translocation (Tat) pathway transport of a precursor protein covalently anchored to its initial cpTatC binding site. J. Biol. Chem. 281, 6130–6135 (2006)

    CAS  Article  Google Scholar 

  17. 17

    Holzapfel, E. et al. The entire N-terminal half of TatC is involved in twin-arginine precursor binding. Biochemistry 46, 2892–2898 (2007)

    CAS  Article  Google Scholar 

  18. 18

    Kreutzenbeck, P. et al. Escherichia coli twin arginine (Tat) mutant translocases possessing relaxed signal peptide recognition specificities. J. Biol. Chem. 282, 7903–7911 (2007)

    CAS  Article  Google Scholar 

  19. 19

    Strauch, E. M. & Georgiou, G. Escherichia coli tatC mutations that suppress defective twin-arginine transporter signal peptides. J. Mol. Biol. 374, 283–291 (2007)

    CAS  Article  Google Scholar 

  20. 20

    Mori, H. & Cline, K. A twin arginine signal peptide and the pH gradient trigger reversible assembly of the thylakoid ΔpH/Tat translocase. J. Cell Biol. 157, 205–210 (2002)

    CAS  Article  Google Scholar 

  21. 21

    Dabney-Smith, C., Mori, H. & Cline, K. Oligomers of Tha4 organize at the thylakoid Tat translocase during protein transport. J. Biol. Chem. 281, 5476–5483 (2006)

    CAS  Article  Google Scholar 

  22. 22

    Cline, K. & McCaffery, M. Evidence for a dynamic and transient pathway through the TAT protein transport machinery. EMBO J. 26, 3039–3049 (2007)

    CAS  Article  Google Scholar 

  23. 23

    Frobel, J., Rose, P. & Muller, M. Early contacts between substrate proteins and TatA translocase component in twin-arginine translocation. J. Biol. Chem. 286, 43679–43689 (2011)

    Article  Google Scholar 

  24. 24

    Fritsch, M. J., Krehenbrink, M., Tarry, M. J., Berks, B. C. & Palmer, T. Processing by rhomboid protease is required for Providencia stuartii TatA to interact with TatC and to form functional homo-oligomeric complexes. Mol. Microbiol. 84, 1108–1123 (2012)

    CAS  Article  Google Scholar 

  25. 25

    Chae, P. S. et al. Maltose-neopentyl glycol (MNG) amphiphiles for solubilization, stabilization and crystallization of membrane proteins. Nature Methods 7, 1003–1008 (2010)

    CAS  Article  Google Scholar 

  26. 26

    Cline, K. & Mori, H. Thylakoid ΔpH-dependent precursor proteins bind to a cpTatC–Hcf106 complex before Tha4-dependent transport. J. Cell Biol. 154, 719–730 (2001)

    CAS  Article  Google Scholar 

  27. 27

    Punginelli, C. et al. Cysteine scanning mutagenesis and topological mapping of the Escherichia coli twin-arginine translocase TatC Component. J. Bacteriol. 189, 5482–5494 (2007)

    CAS  Article  Google Scholar 

  28. 28

    Jeong, K. J. et al. A periplasmic fluorescent reporter protein and its application in high-throughput membrane protein topology analysis. J. Mol. Biol. 341, 901–909 (2004)

    CAS  Article  Google Scholar 

  29. 29

    Behrendt, J., Standar, K., Lindenstrauss, U. & Bruser, T. Topological studies on the twin-arginine translocase component TatC. FEMS Microbiol. Lett. 234, 303–308 (2004)

    CAS  Article  Google Scholar 

  30. 30

    Drew, D. et al. Rapid topology mapping of Escherichia coli inner-membrane proteins by prediction and PhoA/GFP fusion analysis. Proc. Natl Acad. Sci. USA 99, 2690–2695 (2002)

    ADS  CAS  Article  Google Scholar 

  31. 31

    Gouffi, K., Santini, C. L. & Wu, L. F. Topology determination and functional analysis of the Escherichia coli TatC protein. FEBS Lett. 525, 65–70 (2002)

    CAS  Article  Google Scholar 

  32. 32

    Kneuper, H. et al. Molecular dissection of TatC defines critical regions essential for protein transport and a TatB-TatC contact site. Mol. Microbiol. 85, 945–961 (2012)

    CAS  Article  Google Scholar 

  33. 33

    Forrest, L. R., Kramer, R. & Ziegler, C. The structural basis of secondary active transport mechanisms. Biochim. Biophys. Acta 1807, 167–188 (2011)

    CAS  Article  Google Scholar 

  34. 34

    Buchanan, G. et al. Functional complexity of the twin-arginine translocase TatC component revealed by site-directed mutagenesis. Mol. Microbiol. 43, 1457–1470 (2002)

    CAS  Article  Google Scholar 

  35. 35

    Mould, R. M. & Robinson, C. A proton gradient is required for the transport of two lumenal oxygen-evolving proteins across the thylakoid membrane. J. Biol. Chem. 266, 12189–12193 (1991)

    CAS  PubMed  Google Scholar 

  36. 36

    Zoufaly, S. et al. Mapping precursor-binding site on TatC subunit of twin arginine-specific protein translocase by site-specific photo cross-linking. J. Biol. Chem. 287, 13430–13441 (2012)

    CAS  Article  Google Scholar 

  37. 37

    Lausberg, F. et al. Genetic evidence for a tight cooperation of TatB and TatC during productive recognition of twin-arginine (Tat) signal peptides in Escherichia coli. PLoS ONE 7, e39867 (2012)

    ADS  CAS  Article  Google Scholar 

  38. 38

    Bolhuis, A., Mathers, J. E., Thomas, J. D., Barrett, C. M. & Robinson, C. TatB and TatC form a functional and structural unit of the twin-arginine translocase from Escherichia coli. J. Biol. Chem. 276, 20213–20219 (2001)

    CAS  Article  Google Scholar 

  39. 39

    Tarry, M. J. et al. Structural analysis of substrate binding by the TatBC component of the twin-arginine protein transport system. Proc. Natl Acad. Sci. USA 106, 13284–13289 (2009)

    ADS  CAS  Article  Google Scholar 

  40. 40

    Orriss, G. L. et al. TatBC, TatB, and TatC form structurally autonomous units within the twin arginine protein transport system of Escherichia coli. FEBS Lett. 581, 4091–4097 (2007)

    CAS  Article  Google Scholar 

  41. 41

    Maldonado, B., Buchanan, G., Muller, M., Berks, B. C. & Palmer, T. Genetic evidence for a TatC dimer at the core of the Escherichia coli twin arginine (Tat) protein translocase. J. Mol. Microbiol. Biotechnol. 20, 168–175 (2011)

    CAS  Article  Google Scholar 

  42. 42

    Koch, S., Fritsch, M. J., Buchanan, G. & Palmer, T. Escherichia coli TatA and TatB proteins have N-out, C-in topology in intact cells. J. Biol. Chem. 287, 14420–14431 (2012)

    CAS  Article  Google Scholar 

  43. 43

    Greene, N. P. et al. Cysteine scanning mutagenesis and disulfide mapping studies of the TatA component of the bacterial twin arginine translocase. J. Biol. Chem. 282, 23937–23945 (2007)

    CAS  Article  Google Scholar 

  44. 44

    Drew, D., Lerch, M., Kunji, E., Slotboom, D. J. & de Gier, J. W. Optimization of membrane protein overexpression and purification using GFP fusions. Nature Methods 3, 303–313 (2006)

    CAS  Article  Google Scholar 

  45. 45

    Chen, V. B. et al. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D 66, 12–21 (2010)

    CAS  Article  Google Scholar 

  46. 46

    Ashkenazy, H., Erez, E., Martz, E., Pupko, T. & Ben-Tal, N. ConSurf 2010: calculating evolutionary conservation in sequence and structure of proteins and nucleic acids. Nucleic Acids Res. 38, W529–W533 (2010)

    CAS  Article  Google Scholar 

  47. 47

    Tarry, M., Skaar, K., Heijne, G., Draheim, R. R. & Hogbom, M. Production of human tetraspanin proteins in Escherichia coli. Protein Expr. Purif. 82, 373–379 (2012)

    CAS  Article  Google Scholar 

  48. 48

    Deckert, G. et al. The complete genome of the hyperthermophilic bacterium Aquifex aeolicus. Nature 392, 353–358 (1998)

    ADS  CAS  Article  Google Scholar 

  49. 49

    Kawate, T. & Gouaux, E. Fluorescence-detection size-exclusion chromatography for precrystallization screening of integral membrane proteins. Structure 14, 673–681 (2006)

    CAS  Article  Google Scholar 

  50. 50

    Van Duyne, G. D., Standaert, R. F., Karplus, P. A., Schreiber, S. L. & Clardy, J. Atomic structures of the human immunophilin FKBP-12 complexes with FK506 and rapamycin. J. Mol. Biol. 229, 105–124 (1993)

    CAS  Article  Google Scholar 

  51. 51

    Winter, G. xia2: an expert system for macromolecular crystallography data reduction. J. Appl. Cryst. 43, 186–190 (2010)

    CAS  Article  Google Scholar 

  52. 52

    Vonrhein, C., Blanc, E., Roversi, P. & Bricogne, G. Automated structure solution with autoSHARP. Methods Mol. Biol. 364, 215–230 (2007)

    CAS  PubMed  Google Scholar 

  53. 53

    Adams, P. D. et al. The Phenix software for automated determination of macromolecular structures. Methods 55, 94–106 (2011)

    CAS  Article  Google Scholar 

  54. 54

    Bricogne, G., Vonrhein, C., Flensburg, C., Schiltz, M. & Paciorek, W. Generation, representation and flow of phase information in structure determination: recent developments in and around SHARP 2.0. Acta Crystallogr. D 59, 2023–2030 (2003)

    CAS  Article  Google Scholar 

  55. 55

    Abrahams, J. P. & Leslie, A. G. Methods used in the structure determination of bovine mitochondrial F1 ATPase. Acta Crystallogr. D 52, 30–42 (1996)

    CAS  Article  Google Scholar 

  56. 56

    Bricogne, G. et al. BUSTER 2.11.2 (Global Phasing Ltd., 2011)

  57. 57

    Cowtan, K. The Buccaneer software for automated model building. 1. Tracing protein chains. Acta Crystallogr. D 62, 1002–1011 (2006)

    Article  Google Scholar 

  58. 58

    Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D 66, 486–501 (2010)

    CAS  Article  Google Scholar 

  59. 59

    Houtman, J. C. et al. Studying multisite binary and ternary protein interactions by global analysis of isothermal titration calorimetry data in SEDPHAT: application to adaptor protein complexes in cell signaling. Protein Sci. 16, 30–42 (2007)

    CAS  Article  Google Scholar 

  60. 60

    Hess, B., Kutzner, C., van der Spoel, D. & Lindahl, E. GROMACS 4: Algorithms for highly efficient, load-balanced, and scalable molecular simulation. J. Chem. Theory Comput. 4, 435–447 (2008)

    CAS  Article  Google Scholar 

  61. 61

    Monticelli, L., Sorin, E. J., Tieleman, D. P., Pande, V. S. & Colombo, G. Molecular simulation of multistate peptide dynamics: A comparison between microsecond timescale sampling and multiple shorter trajectories. J. Comput. Chem. 29, 1740–1752 (2008)

    CAS  Article  Google Scholar 

  62. 62

    Scott, K. A. et al. Coarse-grained MD simulations of membrane protein-bilayer self-assembly. Structure 16, 621–630 (2008)

    CAS  Article  Google Scholar 

  63. 63

    Monticelli, L., Sorin, E. J., Tieleman, D. P., Pande, V. S. & Colombo, G. Molecular simulation of multistate peptide dynamics: a comparison between microsecond timescale sampling and multiple shorter trajectories. J. Comput. Chem. 29, 1740–1752 (2008)

    CAS  Article  Google Scholar 

  64. 64

    Stansfeld, P. J. & Sansom, M. S. P. From coarse grained to atomistic: a serial multiscale approach to membrane protein simulations. J. Chem. Theory Comput. 7, 1157–1166 (2011)

    CAS  Article  Google Scholar 

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We thank D. Byrne, G. Orriss and R. Owens for their contributions to the early stages of this project; R. Keller for advice; and J. Willem de Gier, D. Daley and R. Huber for providing strains and reagents. This work was supported by the Wellcome Trust (studentships to S.E.R., J.E.G. and M.A.M.; grant 083599, P.R.; grant 092970MA, M.S.P.S.), the Swedish Foundation for Strategic Research (‘Future research leaders 4’ to M.H.), the Swedish Research Council (grant 2010-5061 to M.H.), the E .P. Abrahams Cephalosporin Trust (M.K. and F.R.), the Biotechnology and Biological Sciences Research Council (studentship, M.J.L.; grant BB/E023347/1, S-M.L.; grant BB/1019855/1, P.J.S.), the Medical Research Council (grant G1001640, F.J.; grant G0900888, S.J.), and the European Research Council (Advanced Grant IMPRESS, J.M. and C.V.R.). Work in S.M.L.’s group is funded by the James Martin 21st Century School Vaccine Design Institute.

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The experiments were designed and manuscript written by S.E.R., M.J.T., B.C.B, S.M.L., M.H. and T.P. Experimental work was performed as follows: crystallization trials: S.E.R. and M.J.T.; structure determination: S.E.R., P.R. and S.M.L.; cloning and expression screening: J.E.G., M.J.T., M.J., S-M.L. and M.J.L.; homogeneity screening: M.J.T., M.J., S.E.R., M.A.M. and S.J.; MD simulations: P.J.S. and M.S.P.S; disulphide crosslinking: F.J. and T.P.; signal peptide binding and BN-PAGE: S.E.R., M.K. and F.R.; mass spectrometry: J.M. and C.V.R.

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Correspondence to Ben C. Berks or Susan M. Lea.

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Rollauer, S., Tarry, M., Graham, J. et al. Structure of the TatC core of the twin-arginine protein transport system. Nature 492, 210–214 (2012).

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