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.

  • Brief Communication
  • Published:

Fluorescence fluctuations of quantum-dot sensors capture intracellular protein interaction dynamics

Abstract

We extend the in vitro principle of co-immunoprecipitation to quantify dynamic protein interactions in living cells. Using a multiresolution implementation of fluorescence correlation spectroscopy to achieve maximal temporal resolution, we monitored the interactions of endogenous bait proteins, recruited by quantum dots, with fluorescently tagged prey. With this approach, we analyzed the rapid physiological regulation of protein kinase A.

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

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

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

Figure 1: Quantum dot–based visual immunoprecipitation (QD-VIP).
Figure 2: Optimizing the temporal resolution of QD-VIP by multiparametric analysis and mrFCS.
Figure 3: Monitoring PKA response to physiological stimulations.

Similar content being viewed by others

References

  1. Niethammer, P. et al. PLoS Biol. 5, e29 (2007).

    Article  Google Scholar 

  2. Carlson, C.R. et al. J. Biol. Chem. 281, 21535–21545 (2006).

    Article  CAS  Google Scholar 

  3. Bacia, K., Kim, S.A. & Schwille, P. Nat. Methods 3, 83–89 (2006).

    Article  CAS  Google Scholar 

  4. Lohse, M.J. et al. Trends Pharmacol. Sci. 29, 159–165 (2008).

    Article  CAS  Google Scholar 

  5. Houge, G., Steinberg, R.A., Ogreid, D. & Doskeland, S.O. J. Biol. Chem. 265, 19507–19516 (1990).

    CAS  PubMed  Google Scholar 

  6. Nikolaev, V.O., Gambaryan, S., Engelhardt, S., Walter, U. & Lohse, M.J. J. Biol. Chem. 280, 1716–1719 (2005).

    Article  CAS  Google Scholar 

  7. Violin, J.D. et al. J. Biol. Chem. 283, 2949–2961 (2008).

    Article  CAS  Google Scholar 

  8. Huang, L.J. & Taylor, S.S. J. Biol. Chem. 273, 26739–26746 (1998).

    Article  CAS  Google Scholar 

  9. Seet, B.T., Dikic, I., Zhou, M.M. & Pawson, T. Nat. Rev. Mol. Cell Biol. 7, 473–483 (2006).

    Article  CAS  Google Scholar 

  10. Peyker, A., Rocks, O. & Bastiaens, P.I. ChemBioChem 6, 78–85 (2005).

    Article  CAS  Google Scholar 

  11. Digman, M.A. et al. Biophys. J. 89, 1317–1327 (2005).

    Article  CAS  Google Scholar 

  12. Digman, M.A. et al. Biophys. J. 88, L33–L36 (2005).

    Article  CAS  Google Scholar 

  13. Kolin, D.L. & Wiseman, P.W. Cell Biochem. Biophys. 49, 141–164 (2007).

    Article  CAS  Google Scholar 

  14. Hebert, B., Costantino, S. & Wiseman, P.W. Biophys. J. 88, 3601–3614 (2005).

    Article  CAS  Google Scholar 

  15. Zamir, E. & Bastiaens, P.I. Nat. Chem. Biol. 4, 643–647 (2008).

    Article  CAS  Google Scholar 

  16. Zaccolo, M. et al. Nat. Cell Biol. 2, 25–29 (2000).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by the European Molecular Biology Organization (fellowship to E.Z.) and the Federation of European Biochemical Societies (fellowship to P.H.M.L.). H.E.G. and P.H.M.L. were also supported by the Center for Systems Biology co-financed by European Regional Development Fund and the state of North-Rhine Westphalia. E.Z. was also supported by the EU integrated project grant “Interaction Proteome”. Part of the work was conducted at the Cell Biology and Biophysics Unit, European Molecular Biology Laboratory. We thank M. Zaccolo (Dulbecco Telethon Institute, Venetian Institute of Molecular Medicine, Padova) for the PKA-Cα-EYFP plasmid, S.S. Taylor and R.Y. Tsien (University of California, San Diego) for the mGFP-PKA-RIα plasmid, S. Gentz and C.F. Becker (Max Planck Institute of Molecular Physiology, Dortmund) for the synthesis of the RIAD peptide, M.A. Hink for helpful discussions and A. Krämer for help with manuscript preparation.

Author information

Authors and Affiliations

Authors

Contributions

E.Z., P.H.M.L. and P.I.H.B. devised the method. E.Z. and P.H.M.L. performed the experiments and analyzed the data. E.Z., P.H.M.L., A.K. and H.E.G. developed the analysis. E.Z., P.H.M.L. and P.I.H.B. wrote the paper.

Corresponding author

Correspondence to Philippe I H Bastiaens.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–4, Supplementary Tables 1–2, Supplementary Notes 1–2 (PDF 2918 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Zamir, E., Lommerse, P., Kinkhabwala, A. et al. Fluorescence fluctuations of quantum-dot sensors capture intracellular protein interaction dynamics. Nat Methods 7, 295–298 (2010). https://doi.org/10.1038/nmeth.1441

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nmeth.1441

This article is cited by

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