ABSTRACT
One of the bottlenecks on the route that eventually leads to membrane protein structure through to activity and function is found at the crystal production stage. Diffraction-quality crystals, with which structure is determined, are particularly difficult to prepare currently when a membrane source is used. The reason for this is our limited ability to manipulate proteins with hydrophobic/amphipathic surfaces that are usually enveloped with membrane lipid. More often than not, the protein gets trapped as an intractable aggregate in its watery course from membrane to crystal. As a result, access to the structure and thus function of tens of thousands of membrane proteins is limited. In contrast, a veritable cornucopia of soluble proteins has offered up their structure and valuable insight into function, reflecting the relative ease with which they are crystallized. There exists therefore an enormous need for new ways of producing crystals of membrane proteins. One approach that looks promising involves overexpression as insoluble cytoplasmic inclusion bodies, refolding, and subsequent crystallization (Buchanan, 1999). Another makes use of antibodies to expand polar contacts in the crystal (Ostermeier and Michel, 1997). Very recently, a micelle-based method has been introduced that may share mechanistic similarities with the method described below (Faham and Bowie, 2002).
