Dynamic membrane topology in an unassembled membrane protein

Article metrics

Abstract

Helical membrane proteins are typically assumed to attain stable transmembrane topologies immediately upon co-translational membrane insertion. Here we show that unassembled monomers of the small multidrug resistance (SMR) family exist in a dynamic equilibrium where the N-terminal transmembrane helix flips in and out of the membrane, with rates that depend on dimerization and the polypeptide sequence. Thus, membrane topology can display rapid dynamics in vivo and can be regulated by post-translational assembly.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: Topology of EmrE loops.
Fig. 2: Factors affecting topological dynamics.
Fig. 3: Hydrophobicity of TMHs and loops in the E. coli membrane proteome.

Data availability

The data that support the findings of this study are available from the corresponding authors upon reasonable request.

References

  1. 1.

    Cymer, F., von Heijne, G. & White, S. H. J. Mol. Biol. 427, 999–1022 (2015).

  2. 2.

    Marquette, A. & Bechinger, B. Biomolecules 8, 18 (2018).

  3. 3.

    Bleicken, S. et al. Cell Death Differ. 25, 1717–1731 (2018).

  4. 4.

    Park, T., Struck, D. K., Deaton, J. F. & Young, R. Proc. Natl Acad. Sci. USA 103, 19713–19718 (2006).

  5. 5.

    Buck, T. M., Wagner, J., Grund, S. & Skach, W. R. Nat. Struct. Mol. Biol. 14, 762–769 (2007).

  6. 6.

    Andreev, O. A. et al. Proc. Natl Acad. Sci. USA 107, 4081–4086 (2010).

  7. 7.

    LeBarron, J. & London, E. Langmuir 32, 10752–10760 (2016).

  8. 8.

    Vitrac, H., MacLean, D. M., Jayaraman, V., Bogdanov, M. & Dowhan, W. Proc. Natl Acad. Sci. USA 112, 13874–13879 (2015).

  9. 9.

    Bogdanov, M., Xie, J., Heacock, P. & Dowhan, W. J. Cell Biol. 182, 925–935 (2008).

  10. 10.

    Woodall, N. B., Hadley, S., Yin, Y. & Bowie, J. U. Protein Sci. 26, 824–833 (2017).

  11. 11.

    Woodall, N. B., Yin, Y. & Bowie, J. U. Nat. Commun. 6, 8099 (2015).

  12. 12.

    Fluman, N., Tobiasson, V. & von Heijne, G. Proc. Natl Acad. Sci. USA 114, 7987–7992 (2017).

  13. 13.

    Schuldiner, S. Biochim. Biophys. Acta Proteins Proteom. 1794, 748–762 (2009).

  14. 14.

    Nasie, I., Steiner-Mordoch, S., Gold, A. & Schuldiner, S. J. Biol. Chem. 285, 15234–15244 (2010).

  15. 15.

    Chen, Y.-J. et al. Proc. Natl Acad. Sci. USA 104, 18999–19004 (2007).

  16. 16.

    Rapp, M., Seppälä, S., Granseth, E. & von Heijne, G. Science 315, 1282–1284 (2007).

  17. 17.

    Seppälä, S., Slusky, J. S., Lloris-Garcerá, P., Rapp, M. & von Heijne, G. Science 328, 1698–1700 (2010).

  18. 18.

    Van Lehn, R. C., Zhang, B. & Miller, T. F. Elife 4, e08697 (2015).

  19. 19.

    Bogdanov, M., Zhang, W., Xie, J. & Dowhan, W. Methods 36, 148–171 (2005).

  20. 20.

    Hessa, T. et al. Nature 450, 1026–1030 (2007).

  21. 21.

    Popot, J. L. & Engelman, D. M. Biochemistry 29, 4031–4037 (1990).

  22. 22.

    Schafer, N. P., Truong, H. H., Otzen, D. E., Lindorff-Larsen, K. & Wolynes, P. G. Proc. Natl Acad. Sci. USA 113, 2098–2103 (2016).

  23. 23.

    Min, D., Jefferson, R. E., Bowie, J. U. & Yoon, T.-Y. Nat. Chem. Biol. 11, 981–987 (2015).

  24. 24.

    Lu, W., Schafer, N. P. & Wolynes, P. G. Nat. Commun. 9, 4949 (2018).

  25. 25.

    Feige, M. J. & Hendershot, L. M. Mol. Cell 51, 297–309 (2013).

  26. 26.

    Baba, T. et al. Mol. Syst. Biol. 2, 2006.0008 (2006).

  27. 27.

    Thomason, L. C., Costantino, N. & Court, D. L. E. Curr. Protoc. Mol. Biol. 79, 1.17.1–1.17.8 (2007).

  28. 28.

    Nasie, I., Steiner-Mordoch, S. & Schuldiner, S. Methods Mol. Biol 1033, 121–130 (2013).

  29. 29.

    Odom, O. W., Kudlicki, W., Kramer, G. & Hardesty, B. Anal. Biochem. 245, 249–252 (1997).

  30. 30.

    Tsirigos, K. D., Peters, C., Shu, N., Käll, L. & Elofsson, A. Nucleic Acids Res. 43, W401–W407 (2015).

Download references

Acknowledgements

This work was supported by grants from the Knut and Alice Wallenberg Foundation (2012.0282), the Swedish Research Council (621-2014-3713) and the Swedish Cancer Foundation (15 0888) to G.v.H. N.F. was supported by long-term postdoctoral fellowships from EMBO/Marie Curie Actions (ALTF 211-2014) and from HFSP (LT000277/2015-L).

Author information

M.S. performed and analyzed kinetic studies of monomeric EmrE and mutants and designed the hydrophobicity mutants. M.E. initiated and analyzed the kinetic studies of monomeric EmrE and of several controls. G.v.H. designed the overall study, analyzed data and wrote the paper. N.F. designed the overall study, analyzed data and wrote the paper, performed and analyzed steady state and some kinetic studies of EmrE monomers, performed and analyzed kinetic measurements of SMR and EmrE dimers, developed the kinetic scheme and performed bioinformatic analyses.

Correspondence to Gunnar von Heijne or Nir Fluman.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary Information

Supplementary Figures 1–11

Reporting Summary

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark