Synthesis and Evaluation of Novel Surfactants for Membrane Protein Structural Bio

蛋白质的制备是研究膜蛋白结构和功能的主要瓶颈。目前的结构基因组学研究几乎忽略了这类重要的蛋白质，因为很难生产出足够数量的功能性纯化样品，这是结晶实验所需要的。在膜蛋白样品生产的许多具有挑战性的步骤中，溶解过程？从它们的天然脂质双层中除去疏水膜蛋白并通过使用去污剂胶束人工地使它们与水相相容通常是最成问题的。众所周知，洗涤剂的选择是一种反复试验的经验奋进，已被证明是非常蛋白质特异性的。目前，该领域受到表面活性剂的限制 仅显示窄范围的化学和物理性质的集合。只有少数几类表面活性剂已成功用于约60种独特膜蛋白的纯化和结晶，这些蛋白的结构已保存在蛋白质数据库中。我们寻求开发有效的方法，表面活性剂可以筛选其适用于膜蛋白结构生物学。我们将首先使用表面活性剂测试这些协议，这些表面活性剂在溶解和维持膜蛋白的结构和功能完整性方面具有已证实的实用性。然后，我们将调整这些方案进行高通量筛选，以评估我们的团队从商业实体获得的表面活性剂库。这些表面活性剂是为其他目的而开发的，但其中一些可能适用于这一新兴领域。同时，我们将探索新的合成分子， 旨在替代传统上用于溶解膜蛋白的去污剂。后一类分子的成员被专门设计用于膜蛋白溶解和结晶。

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.