One the primary barriers to membrane protein structure determination by x-ray crystallography is obtaining high quality crystals. We propose two new technologies to improve our ability to obtain crystals of membrane proteins. Method I: We propose a general, high-throughput method to screen for compounds that bind to a target membrane protein. The compounds will stabilize and possibly rigidify the protein, improving the chances of crystallization. The screening method involves a simple test for protein stability. Method II: We proposed to utilize a polymerizing protein module to drive the crystallization of an attached membrane protein. We have demonstrated that the protein module can induce the crystallization of a wide variety of soluble proteins, including those otherwise refractory to crystallization. Moreover, the module is compatible with detergents and preliminary tests indicate that it can drive the crystallization of a transmembrane helix. We plan to further optimize this protein module and test its ability to produce crystals of a range of membrane proteins. Health Relevance: Many diseases are caused by aberrant protein function. Protein function is intimately tied to the elaborate three-dimensional shapes that proteins adopt. It is therefore an important medical goal to learn protein structures so that we can understand how they work and how we can intervene when their functions go awry in disease. This proposal seeks to increase the rate at which we can obtain this critical structural information.
