- Abstract:
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Background: The reconstitution of membrane proteins and complexes into nanoscale lipid bilayer structures has contributed significantly to biochemical and biophysical analyses. Current methods for performing such reconstitutions entail an initial detergent-mediated step to solubilize and isolate membrane proteins. Exposure to detergents, however, can destabilize many membrane proteins and result in a loss of function. Amphipathic copolymers have recently been used to stabilize membrane protei...
Expand abstract - Publication status:
- Published
- Peer review status:
- Peer reviewed
- Version:
- Publisher's version
- Files:
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(bin, 360.2KB)
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(pdf, 1.4MB)
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- Publisher copy:
- 10.1186/1472-6750-13-41
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- Funding agency for:
- Watts, A
- Publisher:
- BioMed Central Ltd. Publisher's website
- Journal:
- BMC Biotechnology Journal website
- Volume:
- 13
- Issue:
- 1
- Pages:
- Article: 41
- Publication date:
- 2013-01-01
- DOI:
- EISSN:
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1472-6750
- ISSN:
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1472-6750
- URN:
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uuid:58200249-f503-4b2f-a746-7103bc816539
- Source identifiers:
-
413522
- Local pid:
- pubs:413522
- Copyright holder:
- Long et al.
- Copyright date:
- 2013
- Notes:
-
176ZE Times Cited:0 Cited References Count:51
Background: The reconstitution of membrane proteins and complexes into nanoscale lipid bilayer structures has contributed significantly to biochemical and biophysical analyses. Current methods for performing such reconstitutions entail an initial detergent-mediated step to solubilize and isolate membrane proteins. Exposure to detergents, however, can destabilize many membrane proteins and result in a loss of function. Amphipathic copolymers have recently been used to stabilize membrane proteins and complexes following suitable detergent extraction. However, the ability of these copolymers to extract proteins directly from native lipid bilayers for subsequent reconstitution and characterization has not been explored. Results: The styrene-maleic acid (SMA) copolymer effectively solubilized membranes of isolated mitochondria and extracted protein complexes. Membrane complexes were reconstituted into polymer-bound nanoscale discs along with endogenous lipids. Using respiratory Complex IV as a model, these particles were shown to maintain the enzymatic activity of multicomponent electron transporting complexes. Conclusions: We report a novel process for reconstituting fully operational protein complexes directly from cellular membranes into nanoscale lipid bilayers using the SMA copolymer. This facile, single-step strategy obviates the requirement for detergents and yields membrane complexes suitable for structural and functional studies.
© 2013 Long et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Journal article
A detergent-free strategy for the reconstitution of active enzyme complexes from native biological membranes into nanoscale discs
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