Section 4.2.6. Membrane-protein crystallization using cubic bicontinuous lipidic phases

Landau & Rosenbusch (1996) introduced the use of bicontinuous cubic phases of lipids for membrane-protein crystallization. In such phases, the lipid forms a single, curved, continuous three-dimensional bilayer [see Lindblom & Rilfors (1989) for a review]. One can incorporate membrane proteins into such a bilayer, as demonstrated with octyl-β-D-glucopyranoside-solubilized monomeric bacteriorhodopsin. The three-dimensional bilayer network serves as a matrix for crystallization. The membrane protein can diffuse through the bilayer, but is also able to establish polar contacts in the third dimension. Landau & Rosenbusch demonstrated that bacteriorhodopsin forms small, well ordered three-dimensional crystals. The X-ray data indicate that the same two-dimensional crystals are present as formed by bacteriorhodopsin in its native environment (the purple membrane). These membranes are now stacked in the third dimension in a well ordered manner. Therefore, these crystals belong to type I.

The method has the conceptual problem that the growing three-dimensional crystal has to disrupt and displace the cubic lipidic phase. Nevertheless, it is hoped that this method can also be used for membrane proteins that do not have a strong tendency to form two-dimensional crystals spontaneously. In particular, this method appears to be the only chance for crystallizing those membrane proteins that are unstable in the absence of added lipids.