Toolkit for High-Resolution Structure and Dynamics of Functional Lipids

项目摘要 膜蛋白在真核细胞中含量丰富，在许多生物中发挥着重要作用。 从细胞黏附和识别到能量产生再到信号级联的各种过程。 此外，膜蛋白约占目前批准的药物靶标的60%， 强调它们与人类疾病的相关性。尽管一些非常高分辨率的结构 膜蛋白现在已经被解决了，我们缺乏也能让我们解决结构的方法 与膜包埋蛋白、外周膜蛋白等相互作用的膜脂 配基。这是尽管特定的膜脂在生物学中发挥着关键的调节作用的事实。我们的术语是 这些“功能性脂”是因为，除了它们在膜上的众所周知的结构作用外，它正在成为 越来越清楚的是，脂类是调节和/或直接执行基本生物功能的效应分子 功能。从原子尺度上理解功能性脂类之间的相互作用是重要的 与人类健康和疾病直接相关的未实现目标。因此，尽管现在有很好的方法来 在非常高的分辨率下解决包括膜蛋白在内的蛋白质的结构，该领域缺乏工具 在高分辨率下解决膜的脂质部分的结构是必要的。这个雄心勃勃的项目旨在 开发一套创新的高分辨率方法“工具包”，供科学界用来解决 调节膜生物功能的脂类结构。我们的方法需要协同和 在整个过程中协调努力：(1)各种磷脂和磷脂的成本效益高、现场特定的同位素标记 (2)将标记的脂类组装成纳米级的脂类双层系统及其生物学上的 相关配体；(3)核磁共振方法，主要是高场魔角 旋转固态核磁共振(SSNMR)，以获得关于与之相互作用的脂类的详细结构信息 配体；(4)使用分子动力学(MD)模拟脂质的尖端计算方法 与双层或双层模拟物中的配体相互作用；以及(5)解决脂结构的新方法。 结合计算核磁共振和MD方法来解决口译中固有的独特挑战 以及理解从包含标记脂质的重复拷贝的平面双层获得的光谱数据 除了它们的特定配体外，还与邻近的脂类相互作用。随着我们研究的进展，我们建议 将此工具包应用于生物学中的典型问题，包括凝血、抗菌肽作用 和类固醇识别。

Project Summary Membrane proteins are abundant in eukaryotic cells and play important roles in a great many biological processes ranging from cell adhesion and recognition to energy production to signaling cascades. Furthermore, membrane proteins make up about 60% of the targets for currently approved drugs, which underscores their relevance to human disease. Although very high resolution structures of a number of membrane proteins have now been solved, we lack methods that will also allow us to resolve the structures of the membrane lipids that interact with membrane-embedded proteins, peripheral membrane proteins and other ligands. This is in spite of the fact that specific membrane lipids play key regulatory roles in biology. We term these “functional lipids” because, in addition to their well-known structural roles in membranes, it is becoming increasingly clear that lipids are effector molecules that modulate and/or directly carry out essential biological functions. An atomic-scale understanding of the interactions carried out by functional lipids is an important unmet goal with direct relevance to human health and disease. Thus, although excellent methods now exist for solving the structures of proteins—including membrane proteins—at very high resolution, the field lacks tools necessary to solve the structures of the lipid part of membranes at high resolution. This ambitious project aims to develop an innovative “toolkit” of high-resolution methods for the scientific community to use in solving the structures of lipids that regulate the biological functions of membranes. Our approach requires synergistic and coordinated efforts throughout: (1) cost-effective, site-specific isotopic labeling of a variety of phospholipids and sterols; (2) assembling labeled lipids into nanoscale lipid bilayer systems together with their biologically relevant ligands; (3) nuclear magnetic resonance (NMR) approaches, principally high-field magic-angle spinning solid-state NMR (SSNMR), to obtain detailed structural information about the lipids interacting with ligands; (4) cutting-edge computational methods employing molecular dynamics (MD) simulations of lipids interacting with ligands in bilayers or bilayer mimetics; and (5) new methods for solving lipid structures by marrying computational NMR and MD approaches to address the unique challenges inherent in interpreting and understanding spectral data obtained from planar bilayers that contain repeating copies of labeled lipids interacting with neighboring lipids in addition to their specific ligands. As our studies progress, we propose to apply this toolkit to exemplary problems in biology, including blood coagulation, antimicrobial peptide action and sterol recognition.