Biological Sciences Division, Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, IL, USA.
Methods Mol Biol. 2021;2187:99-112. doi: 10.1007/978-1-0716-0814-2_6.
Lipid rafts (LRs) represent cellular microdomains enriched in sphingolipids and cholesterol which may fuse to form platforms in which signaling molecules can be organized and regulated (Simons and Ikonen, Nature 387:569-572, 1997; Pike, Biochem J 378:281-292, 2004; Grassme et al., J Immunol 168: 300-307, 2002; Cheng et al., J Exp Med 190:1549-1550, 1999; Kilkus et al., J Neurosci Res 72(1) 62-75, 2003). In a proposed Model 1 (Cheng et al., J Exp Med 190:1549-1550, 1999) the LR has a well-ordered central core composed mainly of cholesterol and sphingolipids that is surrounded by a zone of decreasing lipid order. Detergents such as Triton X-100 can solubilize the core (and a significant amount of phosphoglyceride), but the LRs will be insoluble at 4 °C and be enriched in a well-characterized set of biomarkers. Model 2 proposes that the LRs are homogeneous, but there is selectivity in the lipids (and proteins) extracted by the 1% Triton X-100. Model 3 proposes LRs with distinct lipid compositions are highly structured and can be destroyed by binding molecules such as beta-methylcyclodextrin or filipin. These may be Caveolin in some cell types but not in brain. Since it is unlikely that two LR preparations will be exactly the same this review will concentrate on LRs defined as "small (50 nm) membranous particles which are insoluble in 1% Triton X-100 at 4 °C and have a low buoyant density (Simons and Ikonen, Nature 387:569-572, 1997; Pike, Biochem J 378:281-292, 2004; Grassme et al., J Immunol 168: 300-307, 2002; Cheng et al., J Exp Med 190:1549-1550, 1999; Kilkus et al., J Neurosci Res 72(1):62-75, 2003; Testai et al., J Neurochem 89:636-644, 2004). We will present a generic method for isolating LRs for both lipidomic, proteomic, and cellular signaling analysis [1-6].
脂质筏（LRs）是富含鞘脂和胆固醇的细胞微区，它们可能融合形成平台，在该平台上可以组织和调节信号分子（Simons 和 Ikonen，Nature 387:569-572, 1997； Pike，Biochem J 378:281-292, 2004；Grassme 等人，J Immunol 168: 300-307, 2002；Cheng 等人，J Exp Med 190:1549-1550, 1999；Kilkus 等人，J Neurosci Res 72(1):62-75, 2003）。在提出的模型 1（Cheng 等人，J Exp Med 190:1549-1550, 1999）中，LR 具有一个由胆固醇和鞘脂组成的有序中央核心，周围是脂质有序性降低的区域。去污剂，如 Triton X-100，可以溶解核心（以及大量的磷酸甘油酯），但在 4°C 下，LR 将不溶解，并富含一组特征明确的生物标志物。模型 2 提出 LR 是均匀的，但 1%Triton X-100 提取的脂质（和蛋白质）具有选择性。模型 3 提出具有不同脂质组成的 LR 高度结构化，可以通过结合分子如 beta-甲基环糊精或 filipin 来破坏。这些可能是某些细胞类型中的 Caveolin，但不是在脑中。由于不太可能有两个 LR 制剂完全相同，因此本综述将集中讨论定义为“小（50nm）膜颗粒，在 4°C 下不溶于 1%Triton X-100，且浮力密度低（Simons 和 Ikonen，Nature 387:569-572, 1997； Pike，Biochem J 378:281-292, 2004；Grassme 等人，J Immunol 168: 300-307, 2002；Cheng 等人，J Exp Med 190:1549-1550, 1999；Kilkus 等人，J Neurosci Res 72(1):62-75, 2003；Testai 等人，J Neurochem 89:636-644, 2004）”的 LR。我们将介绍一种用于分离脂质组学、蛋白质组学和细胞信号分析的 LR 的通用方法[1-6]。
