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Fluorescence Correlation Spectroscopy to Examine Protein–Lipid Interactions in Membranes

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Lipid-Protein Interactions

Part of the book series: Methods in Molecular Biology ((MIMB,volume 974))

Abstract

Fluorescence correlation spectroscopy (FCS) is a versatile technique to study membrane dynamics and protein–lipid interactions. It can provide information about diffusion coefficients, concentrations, and molecular interactions of proteins and lipids in the membrane. These parameters allow the determination of protein partitioning into different lipid environments, the identification of lipid domains, and the detection of lipid–protein complexes on the membrane. During the last decade, FCS studies were successfully performed on model membrane systems as also on living cells, to characterize protein–lipid interactions. Recent developments of the method described here improved quantitative measurements on membranes and decreased the number of potential artifacts. The aim of this chapter is to provide the reader with the necessary information and some practical guidelines to perform FCS studies on artificial and cellular membranes.

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References

  1. Bacia K, Schuette CG, Kahya N et al (2004) SNAREs prefer liquid-disordered over “raft” (liquid-ordered) domains when reconstituted into giant unilamellar vesicles. J Biol Chem 279:37951–37955

    Article  PubMed  CAS  Google Scholar 

  2. Doeven MK, Folgering JHA, Krasnikov V et al (2005) Distribution, lateral mobility and function of membrane proteins incorporated into giant unilamellar vesicles. Biophys J 88:1134–1142

    Article  PubMed  CAS  Google Scholar 

  3. Chiantia S, Ries J, Kahya N et al (2006) Combined AFM and two-focus SFCS study of raft-exhibiting model membranes. Chemphyschem 7:2409–2418

    Article  PubMed  CAS  Google Scholar 

  4. Kahya N, Brown DA, Schwille P (2005) Raft partitioning and dynamic behavior of human placental alkaline phosphatase in giant unilamellar vesicles. Biochemistry 44:7479–7489

    Article  PubMed  CAS  Google Scholar 

  5. Takakuwa Y, Pack CG, An XL et al (1999) Fluorescence correlation spectroscopy analysis of the hydrophobic interactions of protein 4.1 with phosphatidyl serine liposomes. Biophys Chem 82:149–155

    Article  PubMed  CAS  Google Scholar 

  6. Ruan Q, Cheng MA, Levi M et al (2004) Spatial–temporal studies of membrane dynamics: scanning fluorescence correlation spectroscopy (SFCS). Biophys J 87:1260–1267

    Article  PubMed  CAS  Google Scholar 

  7. Forstner MB, Yee CK, Parikh AN et al (2006) Lipid lateral mobility and membrane phase structure modulation by protein binding. J Am Chem Soc 128:15221–15227

    Article  PubMed  CAS  Google Scholar 

  8. Kahya N, Wiersma DA, Poolman B et al (2002) Spatial organization of bacteriorhodopsin in model membranes. Light-induced mobility changes. J Biol Chem 277:39304–39311

    Article  PubMed  CAS  Google Scholar 

  9. Liu P, Sudhaharan T, Koh RML et al (2007) Investigation of the dimerization of proteins from the epidermal growth factor receptor family by single wavelength fluorescence cross-correlation spectroscopy. Biophys J 93:684–698

    Article  PubMed  CAS  Google Scholar 

  10. Worch R, Bökel C, Höfinger S et al (2010) Focus on composition and interaction potential of single-pass transmembrane domains. Proteomics 10:4196–4208

    Article  PubMed  CAS  Google Scholar 

  11. García-Sáez AJ, Ries J, Orzáez M et al (2009) Membrane promotes tBID interaction with BCL(XL). Nat Struct Mol Biol 16:1178–1185

    Article  PubMed  Google Scholar 

  12. Honigmann A, Walter C, Erdmann F et al (2010) Characterization of horizontal lipid bilayers as a model system to study lipid phase separation. Biophys J 98:2886–2894

    Article  PubMed  CAS  Google Scholar 

  13. Schwille P, Diez S (2009) Synthetic biology of minimal systems. Crit Rev Biochem Mol Biol 44:223–242

    Article  PubMed  CAS  Google Scholar 

  14. Meacci G, Ries J, Fischer-Friedrich E et al (2006) Mobility of Min-proteins in Escherichia coli measured by fluorescence correlation spectroscopy. Phys Biol 3:255–263

    Article  PubMed  CAS  Google Scholar 

  15. Ries J, Yu SR, Burkhardt M et al (2009) Modular scanning FCS quantifies receptor-ligand interactions in living multicellular organisms. Nat Methods 6:643–645

    Article  PubMed  CAS  Google Scholar 

  16. Schwille P, Korlach J, Webb WW (1999) Fluorescence correlation spectroscopy with single-molecule sensitivity on cell and model membranes. Cytometry 36:176–182

    Article  PubMed  CAS  Google Scholar 

  17. Rajendran L, Schneider A, Schlechtingen G et al (2008) Efficient inhibition of the Alzheimer’s disease beta-secretase by membrane targeting. Science 320:520–523

    Article  PubMed  CAS  Google Scholar 

  18. Cahuzac N, Baum W, Kirkin V et al (2006) Fas ligand is localized to membrane rafts, where it displays increased cell death-inducing activity. Blood 107:2384–2391

    Article  PubMed  CAS  Google Scholar 

  19. Lasserre R, Guo X-J, Conchonaud F et al (2008) Raft nanodomains contribute to Akt/PKB plasma membrane recruitment and activation. Nat Chem Biol 4:538–547

    Article  PubMed  CAS  Google Scholar 

  20. Ries J (2008) Advanced fluorescence correlation techniques to study membrane dynamics. Technical University Dresden

    Google Scholar 

  21. Girard P, Pécréaux J, Lenoir G et al (2004) A new method for the reconstitution of membrane proteins into giant unilamellar vesicles. Biophys J 87:419–429

    Article  PubMed  CAS  Google Scholar 

  22. Kahya N (2010) Light on fluorescent lipids in rafts: a lesson from model membranes. Biochem J 430:e7–e9

    Article  PubMed  CAS  Google Scholar 

  23. Méléard P, Bagatolli LA, Pott T (2009) Giant unilamellar vesicle electroformation from lipid mixtures to native membranes under physiological conditions. Methods Enzymol 465:161–176

    Article  PubMed  Google Scholar 

  24. Dertinger T, Pacheco V, von der Hocht I et al (2007) Two-focus fluorescence correlation spectroscopy: a new tool for accurate and absolute diffusion measurements. Chemphyschem 8:433–443

    Article  PubMed  CAS  Google Scholar 

  25. Korlann Y, Dertinger T, Michalet X et al (2008) Measuring diffusion with polarization–modulation dual-focus fluorescence correlation spectroscopy. Opt Exp 16:14609–14616

    Article  Google Scholar 

  26. Korson L, Drost-Hansen W, Miller FJ (1969) Viscosity of water at various temperatures. J Phys Chem 73:34–39

    Article  CAS  Google Scholar 

  27. Guo L, Har JY, Sankaran J et al (2008) Molecular diffusion measurement in lipid bilayers over wide concentration ranges: a comparative study. Chemphyschem 9:721–728

    Article  PubMed  CAS  Google Scholar 

  28. Machán R, Hof M (2010) Lipid diffusion in planar membranes investigated by fluorescence correlation spectroscopy. Biochim Biophys Acta 1798:1377–1391

    Article  PubMed  Google Scholar 

  29. Ries J, Petrasek Z, Garcia-Saez AJ et al (2010) A comprehensive framework for fluorescence cross-correlation spectroscopy. New J Phys 12:113009–113041

    Article  Google Scholar 

  30. Ries J, Chiantia S, Schwille P (2009) Accurate determination of membrane dynamics with line-scan FCS. Biophys J 96:1999–2008

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

We thank Dr. Fabian Heinemann (BIOTEC, TU Dresden) for the critical reading of this chapter and Erdinc Sezgin (BIOTEC, TU Dresden) for the help with the protocol describing the preparation of living cells for FCS. This work was supported by Dresden International Graduate School for Biomedicine and Bioengineering.

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Authors and Affiliations

  1. Biophysics, BIOTEC, Technische Universitat Dresden, Dresden, Germany

    Viktoria Betaneli & Petra Schwille

Authors
  1. Viktoria Betaneli
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  2. Petra Schwille
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Corresponding author

Correspondence to Petra Schwille .

Editor information

Editors and Affiliations

  1. Institut für Biologie, Abteilung Biophysik, Universität Kassel, Heinrich-Plett-Straße 40, Kassel, 34132, Germany

    Jörg H. Kleinschmidt

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Betaneli, V., Schwille, P. (2013). Fluorescence Correlation Spectroscopy to Examine Protein–Lipid Interactions in Membranes. In: Kleinschmidt, J. (eds) Lipid-Protein Interactions. Methods in Molecular Biology, vol 974. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-275-9_12

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  • DOI: https://doi.org/10.1007/978-1-62703-275-9_12

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-274-2

  • Online ISBN: 978-1-62703-275-9

  • eBook Packages: Springer Protocols

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