Skip to main content

Detergent-Free Membrane Protein Purification

  • Protocol
  • First Online:
Book cover Heterologous Expression of Membrane Proteins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1432))

Abstract

Membrane proteins are localized within a lipid bilayer; in order to purify them for functional and structural studies the first step must involve solubilizing or extracting the protein from these lipids. To date this has been achieved using detergents which disrupt the bilayer and bind to the protein in the transmembrane region. However finding conditions for optimal extraction, without destabilizing protein structure, is time consuming and expensive. Here we present a recently-developed method using a styrene-maleic acid (SMA) co-polymer instead of detergents. The SMA co-polymer extracts membrane proteins in a small disc of lipid bilayer which can be used for affinity chromatography purification, thus enabling the purification of membrane proteins while maintaining their native lipid bilayer environment.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 109.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Gulati S, Jamshad M, Knowles TJ, Morrison KA, Downing R, Cant N, Collins R, Koenderink JB, Ford RC, Overduin M, Kerr ID, Dafforn TR, Rothnie AJ (2014) Detergent-free purification of ABC (ATP-binding-cassette) transporters. Biochem J 461:269–278

    Article  CAS  PubMed  Google Scholar 

  2. Knowles TJ, Finka R, Smith C, Lin YP, Dafforn T, Overduin M (2009) Membrane proteins solubilized intact in lipid containing nanoparticles bounded by styrene maleic acid copolymer. J Am Chem Soc 131:7484–7485

    Article  CAS  PubMed  Google Scholar 

  3. Jamshad M, Charlton J, Lin Y P, Routledge S J, Bawa Z, Knowles T J, Overduin M, Dekker N, Dafforn T R, Bill R M, Poyner D R, Wheatley M (2015) G-protein coupled receptor solubilization and purification for biophysical analysis and functional studies, in the total absence of detergent. Biosci Rep 35(2). pii: e00188

    Google Scholar 

  4. Swainsbury DJ, Scheidelaar S, van Grondelle R, Killian JA, Jones MR (2014) Bacterial reaction centers purified with styrene maleic acid copolymer retain native membrane functional properties and display enhanced stability. Angew Chem Int Ed Engl 53:11803–11807

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Dorr JM, Koorengevel MC, Schafer M, Prokofyev AV, Scheidelaar S, van der Cruijsen EA, Dafforn TR, Baldus M, Killian JA (2014) Detergent-free isolation, characterization, and functional reconstitution of a tetrameric K+ channel: the power of native nanodiscs. Proc Natl Acad Sci U S A 111:18607–18612

    Google Scholar 

  6. Paulin S, Jamshad M, Dafforn TR, Garcia-Lara J, Foster SJ, Galley NF, Roper DI, Rosado H, Taylor PW (2014) Surfactant-free purification of membrane protein complexes from bacteria: application to the staphylococcal penicillin-binding protein complex PBP2/PBP2a. Nanotechnology 25

    Google Scholar 

  7. Postis V, Rawson S, Mitchell JK, Lee SC, Parslow RA, Dafforn TR, Baldwin SA, Muench SP (2015) The use of SMALPs as a novel membrane protein scaffold for structure study by negative stain electron microscopy. Biochim Biophys Acta 1848:496–501

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Long AR, O’Brien CC, Malhotra K, Schwall CT, Albert AD, Watts A, Alder NN (2013) A detergent-free strategy for the reconstitution of active enzyme complexes from native biological membranes into nanoscale discs. BMC Biotechnol 13:41

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Orwick-Rydmark M, Lovett JE, Graziadei A, Lindholm L, Hicks MR, Watts A (2012) Detergent-free incorporation of a seven-transmembrane receptor protein into nanosized bilayer Lipodisq particles for functional and biophysical studies. Nano Lett 12:4687–4692

    Article  CAS  PubMed  Google Scholar 

  10. Jamshad M, Grimard V, Idini I, Knowles TJ, Dowle MR, Schofield N, Sridhar P, Lin YP, Finka R, Wheatley M, Thomas ORT, Palmer RE, Overduin M, Govaerts C, Ruysschaert JM, Edler KJ, Dafforn TR (2015) Structural analysis of a nanoparticle containing a lipid bilayer used for detergent-free extraction of membrane proteins. Nano Res 8:774–789

    Article  CAS  Google Scholar 

  11. Sahu ID, McCarrick RM, Troxel KR, Zhang R, Smith HJ, Dunagan MM, Swartz MS, Rajan PV, Kroncke BM, Sanders CR, Lorigan GA (2013) DEER EPR measurements for membrane protein structures via bifunctional spin labels and lipodisq nanoparticles. Biochemistry 52:6627–6632

    Article  CAS  PubMed  Google Scholar 

  12. Scheidelaar S, Koorengevel MC, Pardo JD, Meeldijk JD, Breukink E, Killian JA (2015) Molecular model for the solubilization of membranes into nanodisks by styrene maleic Acid copolymers. Biophys J 108:279–290

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Alice J. Rothnie .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2016 Springer Science+Business Media New York

About this protocol

Cite this protocol

Rothnie, A.J. (2016). Detergent-Free Membrane Protein Purification. In: Mus-Veteau, I. (eds) Heterologous Expression of Membrane Proteins. Methods in Molecular Biology, vol 1432. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3637-3_16

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-3637-3_16

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3635-9

  • Online ISBN: 978-1-4939-3637-3

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics