Small K+ channels are model proteins for detecting basic interactions between membrane proteins and their surrounding bilayer with relevance for structure and function

只有对蛋白质与其脂质环境之间复杂的相互作用有了深刻的了解，才能完全理解膜蛋白质的结构/功能相关性。在过去的十年里，人们致力于了解跨膜结构域(TMD)的基本结构及其在脂质双层中的嵌入。在此背景下，对简单的合成单膜跨膜蛋白的系统研究为α-螺旋TMDs的氨基酸组成提供了许多基本规则。本项目的目标是扩展这一系统的方法，通过进入更高水平的复杂性来检查膜蛋白的结构/功能相关性，其中包括可测量的功能。这项工作的理想工具是由带有不同TMD的小病毒K+通道提供的。这些功能通道蛋白的一个亚单位是由两个TMD组成的，这两个TMD的长度刚好足以跨越一层膜。从大量的实验和结构工作中，我们已经知道了这些通道的TMDS的基本性质，这对于蛋白质在膜上的锚定是重要的。该项目的成功实现进一步得到了实验进展的支持，该进展允许我们通过在所谓的纳米盘上进行体外翻译来产生通道蛋白。从那里，它们可以在不同类型的脂质双层中重组，它们的功能被记录为单通道波动。在这一坚实的背景下，我们建议系统地研究具有不同TMD的模型通道蛋白在具有不同物理化学特征的脂双层中的功能特性，如可变的双层厚度、不同的头部基团以及有或没有胆固醇。这项工作将得到结构工作的补充，在结构工作中，使用小角度和广角X射线散射方法来确定通道蛋白对脂质双层几何形状的影响。高分辨率显微镜将被用来研究不同膜中通道簇的潜在形成。综上所述，这些互补性的实验将为理解可变脂质环境中简单通道蛋白的结构和功能关系提供坚实的数据基础。我们将获得关于不同膜性质对K+通道功能的影响的详细信息，包括门控和单位电导。这些数据将进一步显示两个伙伴，例如膜和蛋白质，如何在不影响功能的情况下补偿TMD相对于双层厚度的长度不匹配。病毒K+通道的一般结构类似于复杂通道的孔结构，这一事实为将本研究的数据外推到生理上相关的K+通道打开了可能性。

Structure/function correlates of membrane proteins can only be fully understood with profound knowledge on the complex interplay between proteins with their lipid environment. In the last decade much work was devoted to understand the basic architecture of transmembrane domains (TMDs) and their embedding in lipid bilayers. A systematic research with simple synthetic one membrane spanning proteins has in this context provided many of the basic rules for the amino acid composition of a-helical TMDs. The goal of the present project is to extend this systematic approach and examine structure/function correlates of membrane proteins by entering a higher level of complexity, which includes a measurable function. Ideal tools for this endeavor are provided by small viral K+ channels with different TMDs. A subunit of these functional channel proteins is built from two TMDs, which are just long enough to span a membrane. From a bulk of experimental and structural work we already know essential properties of the TMDS of these channels, which are important for an anchoring of the proteins in the membrane. A successful realization of the project is further supported by experimental progress, which allows us to produce channel proteins by in vitro translation in so called nano-discs. From there they can be reconstituted in various types of lipid bilayers and their function recorded as single channel fluctuations. With this solid background we propose to systematically examine the functional properties of model channel proteins with different TMDs in lipid bilayers with distinct physic chemical flavors like variable bilayer thickness, different head groups and with or without cholesterol. This work will be complemented by structural work in which small and wide angle X-ray scattering methods are used to determine the effect of the channel proteins on the geometry of the lipid bilayer. High-resolution microscopy will be employed to examine the potential formation of channel clusters in different membranes. Taken together these complementary experiments will provide a solid data base for an understanding of structure and function relations of simple channel proteins in variable lipid environments. We will obtain detailed information on the influence of different membrane properties on K+ channel function including gating and unitary conductance. The data will further show how the two partners, e.g. the membrane and the protein, compensate mismatches in the length of TMDs with respect to bilayer thickness without compromising function. The fact that the general structure of the viral K+ channels is similar to the pore structure of complex channels opens the possibility to extrapolate data from this research to physiologically relevant K+ channels.