Protein production has been identified as the major bottleneck in the process of analyzing structure and function of membrane proteins. Current structural genomics efforts all but ignore this important class of proteins due to the difficulty in producing sufficient quantities of purified samples in functional form, as required for crystallization experiments. Of the many challenging steps in the production of membrane protein samples, the process of solubilization ? removing the hydrophobic membrane proteins from their native lipid bilayer and artificially rendering them compatible with aqueous phases by the use of detergent micelles - is typically the most problematic. Detergent selection is known to be a trial-and-error, empirical endeavor that has proven to be very protein-specific. Currently, the field is extremely limited by surfactant sets that display only a narrow range of chemical and physical properties. Only a few classes of surfactants have been used successfully in the purification and crystallization of the approximately 60 unique membrane proteins for which structures have been deposited in the Protein Data Bank. We seek to develop efficient methods by which surfactants can be screened for their applicability to membrane protein structural biology. We will initially test these protocols using surfactants with proven utility in solubilization and maintenance of the structural and functional integrity of membrane proteins. We will then adapt these protocols for high-throughput screening in order to assess libraries of surfactants available to our team from commercial entities. These surfactants were developed for other purposes, but some of them might be applicable to this emerging field. In parallel, we will explore novel synthetic molecules that are intended to substitute for the detergents traditionally used for the solubilization of membrane proteins. Members of this latter class of molecules were specifically designed for use in membrane protein solubilization and crystallization.
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