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Crystallizing membrane proteins using lipidic mesophases

Academic Article
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Overview

authors

  • Caffrey, M.
  • Cherezov, Vadim

publication date

  • 2009

journal

  • Nature Protocols  Journal

abstract

  • A detailed protocol for crystallizing membrane proteins that makes use of lipidic mesophases is described. This has variously been referred to as the lipid cubic phase or in meso method. The method has been shown to be quite general in that it has been used to solve X-ray crystallographic structures of prokaryotic and eukaryotic proteins, proteins that are monomeric, homo- and hetero-multimeric, chromophore-containing and chromophore-free, and alpha-helical and beta-barrel proteins. Its most recent successes are the human-engineered beta(2)-adrenergic and adenosine A(2A) G protein-coupled receptors. Protocols are provided for preparing and characterizing the lipidic mesophase, for reconstituting the protein into the monoolein-based mesophase, for functional assay of the protein in the mesophase and for setting up crystallizations in manual mode. Methods for harvesting microcrystals are also described. The time required to prepare the protein-loaded mesophase and to set up a crystallization plate manually is about 1 h.

subject areas

  • Bacterial Outer Membrane Proteins
  • Crystallization
  • Crystallography, X-Ray
  • Escherichia coli Proteins
  • Humans
  • Light-Harvesting Protein Complexes
  • Membrane Lipids
  • Membrane Proteins
  • Membrane Transport Proteins
  • Receptors, Adrenergic, beta-2
  • Receptors, G-Protein-Coupled
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Identity

PubMed Central ID

  • PMC2732203

International Standard Serial Number (ISSN)

  • 1754-2189

Digital Object Identifier (DOI)

  • 10.1038/nprot.2009.31

PubMed ID

  • 19390528
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Additional Document Info

start page

  • 706

end page

  • 731

volume

  • 4

issue

  • 5

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