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Tom40 forms the hydrophilic channel of the mitochondrial import pore for preproteins

Nature volume 395, pages 516–521 (1998)Cite this article

Abstract

The mitochondrial outer membrane contains machinery for the import of preproteins encoded by nuclear genes1,2,3. Eight different Tom (translocase of outer membrane) proteins have been identified that function as receptors and/or are related to a hypothetical general import pore. Many mitochondrial membrane channel activities have been described4,5,6,7, including one related to Tim23 of the inner-membrane protein-import system5; however, the pore-forming subunit(s) of the Tom machinery have not been identified until now. Here we describe the expression and functional reconstitution of Tom40, an integral membrane protein with mainly β-sheet structure. Tom40 forms a cation-selective high-conductance channel that specifically binds to and transports mitochondrial-targeting sequences added to the cis side of the membrane. We conclude that Tom40 is the pore-forming subunit of the mitochondrial general import pore and that it constitutes a hydrophilic, ∼22 Å wide channel for the import of preproteins.

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Figure 1: Expression of yeast Tom40 and its reconstitution into liposomes.
Figure 2: Tom40 forms a high-conductance cation-selective channel.
Figure 3: The Tom40 channel is sensitive to and transports a presequence peptide.

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References

  1. Schatz, G. & Dobberstein, B. Common principles of protein translocation across membranes. Science 271, 1519–1526 (1996).

    Article  ADS  CAS  Google Scholar 

  2. Neupert, W. Protein import into mitochondria. Annu. Rev. Biochem. 66, 863–917 (1997).

    Article  CAS  Google Scholar 

  3. Pfanner, N., Craig, E. A. & Hönlinger, A. Mitochondrial preprotein translocase. Annu. Rev. Cell Dev. Biol. 13, 25–51 (1997).

    Article  CAS  Google Scholar 

  4. Moran, O., Sandri, G., Panfili, E., Stühmer, W. & Sorgato, C. Electrophysiological characterization of contact sites in brain mitochondria. J. Biol. Chem. 265, 908–913 (1990).

    CAS  PubMed  Google Scholar 

  5. Lohret, T. A., Jensen, R. E. & Kinnally, K. W. Tim23, a protein import component of the mitochondrial inner membrane, is required for normal activity of the multiple conductance channel, MCC. J. Cell Biol. 137, 377–386 (1997).

    Article  CAS  Google Scholar 

  6. Juin, P., Thieffry, M., Henry, J.-P. & Vallette, F. M. Relationship between the peptide-sensitive channel and the mitochondrial outer membrane protein translocation machinery. J. Biol. Chem. 272, 6044–6050 (1997).

    Article  CAS  Google Scholar 

  7. Dihanich, M., Schmidt, A., Oppliger, W. & Benz, R. Identification of a new pore in the mitochondrial outer membrane of a porin-deficient yeast mutant. Eur. J. Biochem. 181, 703–708 (1989).

    Article  CAS  Google Scholar 

  8. Baker, K. P., Schaniel, A., Vestweber, D. & Schatz, G. Ayeast mitochondrial outer membrane protein essential for protein import and cell viability. Nature 348, 605–609 (1990).

    Article  ADS  CAS  Google Scholar 

  9. Mannella, C. A., Neuwald, A. F. & Lawrence, C. E. Detection of likely transmembrane β-strand regions in sequences of mitochondrial pore proteins using the Gibbs sampler. J. Bioenerg. Biomembr. 28, 163–169 (1996).

    Article  CAS  Google Scholar 

  10. Schmid, B., Krömer, M. & Schulz, G. E. Expression of porin from Rhodopseudomonas blastica in Escherichia coli inclusion bodies and folding into exact native structure. FEBS Lett. 381, 111–114 (1996).

    Article  CAS  Google Scholar 

  11. Sreerama, N. & Woody, R. W. Protein secondary structure from circular dichroism spectroscopy: combining variable selection principle and cluster analysis with neural network, ridge regression and self-consistent methods. J. Mol. Biol. 242, 497–507 (1994).

    CAS  PubMed  Google Scholar 

  12. Hille, B. in Ionic Channels of Excitable Membranes. Ch. 9, 11 (Sinauer, Sunderland, Massachusetts, 1992).

    Google Scholar 

  13. Hinnah, S., Hill, K., Wagner, R., Schlicher, T. & Soll, J. Reconstitution of a chloroplast protein import channel. EMBO J. 16, 7351–7360 (1997).

    Article  CAS  Google Scholar 

  14. Smart, O. S., Breed, J., Smith, G. R. & Sansom, M. S. Anovel method for structure-based prediction of ion channel conductance properties. Biophys. J. 72, 1109–1126 (1997).

    Article  ADS  CAS  Google Scholar 

  15. Krasilnikov, O. V., Sabirov, R. Z., Ternovsky, V. L., Merzliak, P. G. & Muratkhodjaev, J. N. Asimple method for the determination of the pore radius of ion channels in planar lipid bilayer membranes. FEMS Microbiol. Immunol. 5, 93–100 (1992).

    Article  CAS  Google Scholar 

  16. Alconada, A., Gärtner, F., Hönlinger, A., Kübrich, M. & Pfanner, N. Mitochondrial receptor complex from Neurospora crassa and Saccharomyces cerevisiae. Methods Enzymol. 260, 263–286 (1995).

    Article  CAS  Google Scholar 

  17. Dietmeier, K. et al. Tom5 functionally links mitochondrial preprotein receptors to the general import pore. Nature 388, 195–200 (1997).

    Article  ADS  CAS  Google Scholar 

  18. Dekker, P. J. T. et al. The Tim core complex defines the number of mitochondrial translocation contact sites and can hold arrested preproteins in the absence of matrix Hsp70-Tim44. EMBO J. 16, 5408–5419 (1997).

    Article  CAS  Google Scholar 

  19. Allison, D. S. & Schatz, G. Artificial mitochondrial presequences. Proc. Natl Acad. Sci. USA 83, 9011–9015 (1986).

    Article  ADS  CAS  Google Scholar 

  20. Brix, J., Dietmeier, K. & Pfanner, N. Differential recognition of preproteins by the purified cytosolic domains of the mitochondrial import receptors Tom20, Tom22, and Tom70. J. Biol. Chem. 272, 20730–20735 (1997).

    Article  CAS  Google Scholar 

  21. Rapaport, D., Neupert, W. & Lill, R. Mitochondrial protein import: Tom40 plays a major role in targeting and translocation of preproteins by forming a specific binding site for the presequence. J. Biol. Chem. 272, 18725–18731 (1997).

    Article  CAS  Google Scholar 

  22. Mayer, A., Neupert, W. & Lill, R. Mitochondrial protein import: reversible binding of the presequence at the trans side of the outer membrane drives partial translocation and unfolding. Cell 80, 127–137 (1995).

    Article  CAS  Google Scholar 

  23. Vestweber, D. & Schatz, G. DNA-protein conjugates can enter mitochondria via the protein import pathway. Nature 338, 170–172 (1989).

    Article  ADS  CAS  Google Scholar 

  24. Künkele, K.-P. et al. The preprotein translocation channel of the outer membrane of mitochondria. Cell 93, 1009–1019 (1998).

    Article  Google Scholar 

  25. Matlack, K. E. S., Mothes, W. & Rapoport, T. A. Protein translocation: tunnel vision. Cell 92, 381–390 (1998).

    Article  CAS  Google Scholar 

  26. Vestweber, D., Brunner, J., Baker, A. & Schatz, G. A24K outer-membrane protein is a component of the yeast mitochondrial protein import site. Nature 341, 205–209 (1989).

    Article  ADS  CAS  Google Scholar 

  27. Kiebler, M. et al. Identification of a mitochondrial receptor complex required for recognition and membrane insertion of precursor proteins. Nature 348, 610–616 (1990).

    Article  ADS  CAS  Google Scholar 

  28. Söllner, T. et al. Mapping of the protein import machinery in the mitochondrial outer membrane by crosslinking of translocation intermediates. Nature 355, 84–87 (1992).

    Article  ADS  Google Scholar 

  29. Hönlinger, A. et al. The mitochondrial receptor complex: Mom22 is essential for cell viability and directly interacts with preproteins. Mol. Cell. Biol. 15, 3382–3389 (1995).

    Article  Google Scholar 

  30. Bolliger, L., Junne, T., Schatz, G. & Lithgow, T. Acidic receptor domains on both sides of the outer membrane mediate translocation of precursor proteins into yeast mitochondria. EMBO J. 14, 6318–6326 (1995).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank P. J. T. Dekker, B. Guiard, M. Kiebler, W. Kühlbrandt, D. Mills, J. Brix, S. C. Hinnah and H. Martin for materials, advice and discussion. This work was supported by the Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 388, the Fonds der Chemischen Industrie (N.P.), the Sonderforschungsbereich 171 (R.W.) and a longterm fellowship of the Alexander-von-Humboldt Foundation (M.T.R.)

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Authors and Affiliations

  1. Biophysik, Universität Osnabrück, FB Biologie/Chemie, D-49034, Osnabrück, Germany

    Kerstin Hill & Richard Wagner

  2. Institut für Biochemie und Molekularbiologie, Universität Freiburg, Hermann-Herder-Strasse 7, D-79104, Freiburg, Germany

    Kirstin Model, Michael T. Ryan, Klaus Dietmeier, Falk Martin & Nikolaus Pfanner

  3. Fakultät für Biologie, Universität Freiburg, D-79104, Freiburg, Germany

    Falk Martin

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  1. Kerstin Hill
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Correspondence to Nikolaus Pfanner.

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Hill, K., Model, K., Ryan, M. et al. Tom40 forms the hydrophilic channel of the mitochondrial import pore for preproteins. Nature 395, 516–521 (1998). https://doi.org/10.1038/26780

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