Lipidic sponge phase crystallization of membrane proteins

J Mol Biol. 2006 Nov 17;364(1):44-53. doi: 10.1016/j.jmb.2006.06.043. Epub 2006 Jul 7.

Abstract

Bicontinuous lipidic cubic phases can be used as a host for growing crystals of membrane proteins. Since the cubic phase is stiff, handling is difficult and time-consuming. Moreover, the conventional cubic phase may interfere with the hydrophilic domains of membrane proteins due to the limited size of the aqueous pores. Here, we introduce a new crystallization method that makes use of a liquid analogue of the cubic phase, the sponge phase. This phase facilitates a considerable increase in the allowed size of aqueous domains of membrane proteins, and is easily generalised to a conventional vapour diffusion crystallisation experiment, including the use of nanoliter drop crystallization robots. The appearance of the sponge phase was confirmed by visual inspection, small-angle X-ray scattering and NMR spectroscopy. Crystals of the reaction centre from Rhodobacter sphaeroides were obtained by a conventional hanging-drop experiment, were harvested directly without the addition of lipase or cryoprotectant, and the structure was refined to 2.2 Angstroms resolution. In contrast to our earlier lipidic cubic phase reaction centre structure, the mobile ubiquinone could be built and refined. The practical advantages of the sponge phase make it a potent tool for crystallization of membrane proteins.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • Crystallization
  • Lipids / chemistry*
  • Membrane Proteins / chemistry*
  • Molecular Structure
  • Photosynthetic Reaction Center Complex Proteins / chemistry*
  • Protein Conformation
  • Rhodobacter sphaeroides / chemistry
  • X-Ray Diffraction

Substances

  • Lipids
  • Membrane Proteins
  • Photosynthetic Reaction Center Complex Proteins

Associated data

  • PDB/2GNU