Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Coupling ion channels to receptors for biomolecule sensing

Nature Nanotechnology volume 3, pages 620–625 (2008)Cite this article

Abstract

Nanoscale electrical biosensors are promising tools for diagnostics and high-throughput screening systems. The electrical signal allows label-free assays with a high signal-to-noise ratio and fast real-time measurements. The challenge in developing such biosensors lies in functionally connecting a molecule detector to an electrical switch. Advances in this field have relied on synthetic ion-conducting pores1 and modified ion channels2,3,4 that are not yet suitable for biomolecule screening. Here we report the design and characterization of a novel bioelectric-sensing platform engineered by coupling an ion channel, which serves as the electrical probe, to G-protein-coupled receptors (GPCRs), a family of receptors that detect molecules outside the cell. These ion-channel-coupled receptors may potentially detect a wide range of ligands recognized by natural or altered GPCRs5,6, which are known to be major pharmaceutical targets7. This could form a unique platform for label-free drug screening8.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Principle of ion-channel-coupled receptors (ICCRs).
Figure 2: Design and expression of fusion proteins.
Figure 3: Receptor–channel coupling in M2-based ICCRs.
Figure 4: D2-based ion-channel-coupled receptors.
Figure 5: The fused D2 receptor remains functional.

Similar content being viewed by others

References

  1. Bayley, H. & Jayasinghe, L. Functional engineered channels and pores. Mol. Membr. Biol. 21, 209–220 (2004).

    Article  CAS  Google Scholar 

  2. Bouzat, C. et al. Coupling of agonist binding to channel gating in an ACh-binding protein linked to an ion channel. Nature 430, 896–900 (2004).

    Article  CAS  Google Scholar 

  3. Grutter, T. et al. A chimera encoding the fusion of an acetylcholine-binding protein to an ion channel is stabilized in a state close to the desensitized form of ligand-gated ion channels. CR Biol. 328, 223–234 (2005).

    Article  CAS  Google Scholar 

  4. Ohndorf, U. M. & MacKinnon, R. Construction of a cyclic nucleotide-gated KcsA K+ channel. J. Mol. Biol. 350, 857–865 (2005).

    Article  CAS  Google Scholar 

  5. Kristiansen, K. Molecular mechanisms of ligand binding, signaling, and regulation within the superfamily of G-protein-coupled receptors: molecular modeling and mutagenesis approaches to receptor structure and function. Pharmacol. Ther. 103, 21–80 (2004).

    Article  CAS  Google Scholar 

  6. Armbruster, B. N., Li, X., Pausch, M. H., Herlitze, S. & Roth, B. L. Evolving the lock to fit the key to create a family of G protein-coupled receptors potently activated by an inert ligand. Proc. Natl Acad. Sci. USA 104, 5163–5168 (2007).

    Article  Google Scholar 

  7. Schlyer, S. & Horuk, R. I want a new drug: G-protein-coupled receptors in drug development. Drug Discov. Today 11, 481–493 (2006).

    Article  CAS  Google Scholar 

  8. Thomsen, W., Frazer, J. & Unett, D. Functional assays for screening GPCR targets. Curr. Opin. Biotechnol. 16, 655–665 (2005).

    CAS  Google Scholar 

  9. Moreau, C. J., Prost, A. L., Derand, R. & Vivaudou, M. SUR, ABC proteins targeted by KATP channel openers. J. Mol. Cell. Cardiol. 38, 951–963 (2005).

    Article  CAS  Google Scholar 

  10. Moreau, C. J., Jacquet, H., Prost, A. L., D'Hahan, N. & Vivaudou, M. The molecular basis of the specificity of action of KATP channel openers. EMBO J. 19, 6644–6651 (2000).

    Article  CAS  Google Scholar 

  11. Dupuis, J. P., Revilloud, J., Moreau, C. J. & Vivaudou, M. Three C-terminal residues from the sulphonylurea receptor contribute to the functional coupling between the KATP channel subunits SUR2A and Kir6.2. J. Physiol. 586, 3075–3085 (2008).

    Article  CAS  Google Scholar 

  12. Haider, S., Antcliff, J. F., Proks, P., Sansom, M. S. & Ashcroft, F. M. Focus on Kir6.2: a key component of the ATP-sensitive potassium channel. J. Mol. Cell. Cardiol. 38, 927–936 (2005).

    Article  CAS  Google Scholar 

  13. Nichols, C. G. KATP channels as molecular sensors of cellular metabolism. Nature 440, 470–476 (2006).

    Article  CAS  Google Scholar 

  14. Tucker, S. J., Gribble, F. M., Zhao, C., Trapp, S. & Ashcroft, F. M. Truncation of Kir6.2 produces ATP-sensitive K+ channels in the absence of the sulphonylurea receptor. Nature 387, 179–183 (1997).

    Article  CAS  Google Scholar 

  15. Zerangue, N., Schwappach, B., Jan, Y. N. & Jan, L. Y. A new ER trafficking signal regulates the subunit stoichiometry of plasma membrane KATP channels. Neuron 22, 537–548 (1999).

    Article  CAS  Google Scholar 

  16. Koster, J. C., Sha, Q., Shyng, S. L. & Nichols, C. G. ATP inhibition of KATP channels: control of nucleotide sensitivity by the N-terminal domain of the Kir6.2 subunit. J. Physiol. 515, 19–30 (1999).

    Article  CAS  Google Scholar 

  17. Tucker, S. J. & Ashcroft, F. M. Mapping of the physical interaction between the intracellular domains of an inwardly rectifying potassium channel, Kir6.2. J. Biol. Chem. 274, 33393–33397 (1999).

    Article  CAS  Google Scholar 

  18. Vivaudou, M. et al. Probing the G-protein regulation of GIRK1 and GIRK4, the two subunits of the K-ACh channel, using functional homomeric mutants. J. Biol. Chem. 272, 31553–31560 (1997).

    Article  CAS  Google Scholar 

  19. McKinney, M., Miller, J. H., Gibson, V. A., Nickelson, L. & Aksoy, S. Interactions of agonists with M2 and M4 muscarinic receptor subtypes mediating cyclic AMP inhibition. Mol. Pharmacol. 40, 1014–1022 (1991).

    CAS  Google Scholar 

  20. Beguin, P., Nagashima, K., Nishimura, M., Gonoi, T. & Seino, S. PKA-mediated phosphorylation of the human KATP channel: separate roles of Kir6.2 and SUR1 subunit phosphorylation. EMBO J. 18, 4722–4732 (1999).

    Article  CAS  Google Scholar 

  21. Light, P. E., Bladen, C., Winkfein, R. J., Walsh, M. P. & French, R. J. Molecular basis of protein kinase C-induced activation of ATP-sensitive potassium channels. Proc. Natl Acad. Sci. USA 97, 9058–9063 (2000).

    Article  CAS  Google Scholar 

  22. Wess, J. G-protein-coupled receptors: molecular mechanisms involved in receptor activation and selectivity of G-protein recognition. FASEB J. 11, 346–354 (1997).

    Article  CAS  Google Scholar 

  23. Li, J. H. et al. Distinct structural changes in a G protein-coupled receptor caused by different classes of agonist ligands. J. Biol. Chem. 282, 26284–26293 (2007).

    Article  CAS  Google Scholar 

  24. Kuo, A. L., Domene, C., Johnson, L. N., Doyle, D. A. & Venien-Bryan, C. Two different conformational states of the KirBac3.1 potassium channel revealed by electron crystallography. Structure 13, 1463–1472 (2005).

    Article  CAS  Google Scholar 

  25. Nishida, M., Cadene, M., Chait, B. T. & MacKinnon, R. Crystal structure of a Kir3.1–prokaryotic Kir channel chimera. EMBO J. 26, 4005–4015 (2007).

    Article  CAS  Google Scholar 

  26. Long, S. B., Tao, X., Campbell, E. B. & MacKinnon, R. Atomic structure of a voltage-dependent K+ channel in a lipid membrane-like environment. Nature 450, 376–382 (2007).

    Article  CAS  Google Scholar 

  27. Leifert, W. R., Aloia, A. L., Bucco, O., Glatz, R. V. & McMurchie, E. J. G-protein-coupled receptors in drug discovery: nanosizing using cell-free technologies and molecular biology approaches. J. Biomol. Screen. 10, 765–779 (2005).

    Article  CAS  Google Scholar 

  28. Hosy, E., Derand, R., Revilloud, J. & Vivaudou, M. Remodelling of the SUR–Kir6.2 interface of the KATP channel upon ATP binding revealed by the conformational blocker rhodamine 123. J. Physiol. 582, 27–39 (2007).

    Article  CAS  Google Scholar 

  29. Makarova, O., Kamberov, E. & Margolis, B. Generation of deletion and point mutations with one primer in a single cloning step. Biotechniques 29, 970–972 (2000).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We are grateful to E. Reuveny for PTX–S1, S. Seino for Kir6.2, D. Logothetis for M2, B. Kobilka and R. Stevens for prepublication β2 pdb files, and A. Dupuis, F. Fieschi and N. Hussy for useful discussions. A.-L. Prost and R. Derand with J.R. performed exploratory experiments on non-GPCR fusion proteins that helped guide our research strategy. S. Crouzy and S. Redon helped with molecular dynamics. This project, part of the European project Receptronics coordinated by M. Tartagni, and a postdoctoral fellowship to C.M. were funded by the Sixth Framework Program of the European Union (STREP NMP4-CT-2005-017114). Additional funds were provided by CEA (Commissariat à l'Energie Atomique; Programme Toxicologie Nucléaire Environnementale), CNRS and Université Joseph Fourier. J.D. was supported by a CEA doctoral studentship.

Author information

Authors and Affiliations

  1. Laboratoire des Protéines Membranaires, Institut de Biologie Structurale (CEA, CNRS, UJF), 41 rue Jules Horowitz, Grenoble, 38027, France

    Christophe J. Moreau, Julien P. Dupuis, Jean Revilloud & Michel Vivaudou

  2. Laboratoire Chimie et Biologie des Métaux, Institut de Recherches en Technologies et Sciences pour le Vivant, CEA/Grenoble, 17 rue des Martyrs, Grenoble, 38041, France

    Karthik Arumugam

Authors
  1. Christophe J. Moreau
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Julien P. Dupuis
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jean Revilloud
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Karthik Arumugam
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Michel Vivaudou
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

C.J.M. and M.V. established the concepts of this study, analysed data and wrote the manuscript. C.J.M. performed the experiments with help from J.P.D. (TEVC) and J.R. (molecular biology, TEVC, outside-out recordings). K.A. helped with molecular modelling. M.V. supervised the project.

Corresponding authors

Correspondence to Christophe J. Moreau or Michel Vivaudou.

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Cite this article

Moreau, C., Dupuis, J., Revilloud, J. et al. Coupling ion channels to receptors for biomolecule sensing. Nature Nanotech 3, 620–625 (2008). https://doi.org/10.1038/nnano.2008.242

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nnano.2008.242

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing