Domains in Biologically Relevant Lipid Membranes

生物学相关脂质膜中的结构域

• 批准号: RGPIN-2019-07229
• 负责人: Thewalt, Jenifer
• 金额: $ 2.62万
• 依托单位: Simon Fraser University
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2019
• 资助国家: 加拿大
• 起止时间: 2019-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=686602
• 关键词: Domains; Biologically; Relevant; Lipid; Membranes

Cell membrane structure and dynamics are key to maintaining a properly functioning cell, and lipids have a dominant role in determining the organization of cell membranes. Since the “fluid mosaic” model was published in 1972, awareness has been growing that the "mosaic" aspect of membrane structure is as important as the "fluid" aspect. Integral membrane proteins, particularly those that link to the cytoskeleton and those that can form signalling platforms, certainly determine aspects of the mosaic. Other important mosaic characteristics arise because lipids in membranes spontaneously interact and can congregate in dynamic clusters ranging from nanoscale fluctuations to micron scale phase separation. Small short lived clusters known as "rafts" in cell membranes are challenging to study in vivo. In contrast to cell membranes especially eukaryotic membranes lipid membranes are tractable research subjects. Understanding the physical and chemical interactions among lipids in a membrane with known composition helps to define the playing field that biology can use. Some of the most influential concepts in the study of cell membrane organization originated with investigations into the phase behavior of such "simple" systems. One extremely useful concept is the cholesterol rich liquid ordered (lo) phase. In the lo phase cholesterol damps out lipid chain fluctuations while maintaining rotational and lateral lipid mobility, leading to a more tightly packed and therefore thicker, but still liquid crystalline, membrane.***We map out the fundamental interactions between phospholipids, sterols and sphingolipids in model membranes. Deuterium nuclear magnetic resonance (NMR) is a powerful experimental approach to the study of phase coexistence in membranes; different phases result in clearly distinguishable spectra, and the quantity of labeled lipid in a particular phase can be measured unambiguously. We have concentrated on measuring the phase diagrams of binary lipid mixtures but are now extending our work to ternary mixtures (eg POPC, DPPC or sphingomyelin, sterol) and also ternary mixtures + fluorescent probe. Cell membranes entering apoptosis are postulated to contain gel domains formed as sphingomyelin is converted to ceramide. We will test whether gel phases form in sphingomyelin + ceramide in the presence of the high cholesterol content found in plasma membranes. We will then study an "outer leaflet" plasma membrane model containing POPC, sphingomyelin and cholesterol, then add ceramide to this system.***Interestingly, lipid phase behavior is important to the efficacy of lipid nanoparticles (LNPs) used for drug delivery. With the Cullis and Tieleman research groups, we are mapping the phase propensities of lipids used to manufacture LNPs. We plan to combine NMR and X-ray scattering to gain insight into nonbilayer phases formed by these lipids in the presence of anionic lipids typical of endosomal membranes.**

细胞膜结构和动力学是维持细胞正常功能的关键，而脂质在决定细胞膜的组织方面起着主导作用。自 1972 年“流体马赛克”模型发布以来，人们越来越意识到膜结构的“马赛克”方面与“流体”方面同样重要。整合膜蛋白，特别是那些连接到细胞骨架的蛋白和那些可以形成信号平台的蛋白，无疑决定了嵌合体的各个方面。其他重要的镶嵌特征的出现是因为膜中的脂质自发地相互作用，并且可以聚集在从纳米级波动到微米级相分离的动态簇中。细胞膜中被称为“筏”的短寿命小簇在体内研究具有挑战性。与细胞膜（尤其是真核细胞膜）相比，脂质膜是易于处理的研究对象。了解具有已知成分的膜中脂质之间的物理和化学相互作用有助于定义生物学可以使用的竞争环境。细胞膜组织研究中一些最有影响力的概念源于对此类“简单”系统相行为的研究。一个非常有用的概念是富含胆固醇的液体有序（lo）相。在 lo 相，胆固醇抑制脂质链波动，同时保持旋转和横向脂质流动性，从而形成更紧密的堆积，因此更厚，但仍然是液晶膜。***我们绘制了模型膜中磷脂、甾醇和鞘脂之间的基本相互作用。氘核磁共振 (NMR) 是研究膜中相共存的一种强大的实验方法；不同的相会产生清晰可辨的光谱，并且可以明确地测量特定相中标记的脂质的数量。我们专注于测量二元脂质混合物的相图，但现在将我们的工作扩展到三元混合物（例如POPC、DPPC或鞘磷脂、甾醇）以及三元混合物+荧光探针。假定进入细胞凋亡的细胞膜含有鞘磷脂转化为神经酰胺时形成的凝胶结构域。我们将测试在质膜中存在高胆固醇含量的情况下，鞘磷脂 + 神经酰胺是否会形成凝胶相。然后，我们将研究含有 POPC、鞘磷脂和胆固醇的“外叶”质膜模型，然后将神经酰胺添加到该系统中。***有趣的是，脂质相行为对于用于药物输送的脂质纳米颗粒 (LNP) 的功效非常重要。我们与 Cullis 和 Tieleman 研究小组一起绘制了用于制造 LNP 的脂质的相倾向。我们计划将 NMR 和 X 射线散射相结合，以深入了解这些脂质在内体膜典型的阴离子脂质存在的情况下形成的非双层相。**