Crystallization of membrane proteins in cubo.

Our understanding of lipidic cubic phases for the crystallization of membrane proteins has advanced greatly since the inception of the concept in 1996, and the method is becoming well accepted. Several protocols that allow the efficient screening of crystallization conditions and handling of crystals are presented. State-of-the art micro techniques allow a large number of crystallization conditions to be tested using very small amounts of protein, and diffraction quality crystals can be grown in larger volumes in glass vials. In cubo crystallization conditions differ from those employed for detergent-solubilized proteins. Variations comprise the type of lipid matrix, detergent, protein, salt, temperature, hydration, pH, and pressure. Commercially available screening kits may be applied in order to define lead conditions. Once obtained, crystals may be removed from the surrounding cubic phase mechanically, by enzymatic hydrolysis, or by detergent solubilization. We anticipate this set of protocols to be applied successfully to larger, less stable, and noncolored membrane proteins in order to obtain well-diffracting crystals of membrane proteins that have so far evaded crystallization in the detergent-solubilized state.

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