Development of Parallel-Aligning Magnetically Orientable Model Membranes for Use in NMR Studies of Membrane Proteins

膜蛋白存在于所有生物系统中包裹细胞和亚细胞区室的薄鞘中。这些蛋白质代表了一类重要的生物分子，它们在分子水平上的三维图像或“结构”信息很难获得。原则上，一种被称为“定向样品核磁共振”（OSNMR）的物理技术能够产生实验数据，这些数据可以直接导致膜样环境中膜蛋白的结构。然而，目前OSNMR技术的局限性之一是缺乏一种理想的方法来均匀定向膜蛋白，从而使样品中的所有蛋白质在同一方向上排列。本项目的工作将寻求通过合理设计一类新的膜模拟“模型膜”来克服这一限制，其中纯化的膜蛋白可以被溶解并定向用于OSNMR研究。作为本方案目标的新型膜系统必须同时满足四个标准：(i)水必须是溶剂。（ii）液晶组件必须是可溶的胶束状盘状双层颗粒（“双束”）。（三）单胞体在强磁场作用下必须呈现向列有序，使其双层法线与施加磁场的方向一致。（iv）液晶系统不能使溶解在新介质中的膜蛋白变性。各种因素预测，这些目标将通过涉及一系列新型洗涤剂样分子的液晶来实现：“PF-DAPA”表面活性剂。这些化合物将通过长链全氟羧酸与2,3二氨基丙酸的3-氨基的酰胺缩合而合成。它们的液晶性质将用核磁共振来表征。将测试满足上述第i-iii项标准的组合物，以测试其溶解膜蛋白和使膜蛋白在天然构象状态下具有磁性定向的能力。＊＊＊

Sanders 9513357 Membrane proteins reside in the thin sheaths which encapsulate cells and subcellular compartments throughout all biological systems. These proteins represent an important class of biological molecules for which three-dimensional pictorial or "structural" information on the molecular level has been very difficult to obtain. In principle, a physical technique known as "oriented sample nuclear magnetic resonance" (OSNMR) is capable of yielding experimental data which can lead directly to the structures of membrane proteins in a membrane-like environment. However, one of the limitations of current OSNMR technique is the lack of an ideal method for uniformly orienting membrane proteins so that all of the proteins in a sample are lined up in the same direction. The work of this project will seek to overcome this limitation by carrying out the rational design of a new class of membrane-mimicking "model membranes" in which purified membrane proteins can be solubilized and oriented for OSNMR studies. The novel model membrane system which is the goal of this proposal must simultaneously satisfy four criteria: (i) Water must be the solvent. (ii) The liquid crystalline assemblies must be soluble, micelle-like discoidal bilayered particles ("bicelles"). (iii) The bicelles must exhibit nematic order under the influence of a strong magnetic field such that their bilayer normals align in the direction of the applied field. (iv) The liquid crystalline system must not denature membrane proteins solubilized in the novel media. %%% Various considerations predict that these goals will be met by liquid crystals involving a novel series of detergent-like molecules: the "PF-DAPA" surfactants. These compounds will be synthesized via amide condensation of long chain perfluorinated carboxylic acids with the 3-amino group of 2,3 diaminopropionic acid. Their liquid crystalline properties will be characterized using nuclear magnetic resonance. Compositions which satisfy criteria i-iii above will be tested for their ability to solubilize and magnetically orient membrane proteins in native-like conformational states. ***