Delaunay Jean-Louis, Breton Michelyne, Trugnan Germain, Maurice Michèle
Institut National de la Santé et de la Recherche Médicale UMR S 538, Université Pierre et Marie Curie, Paris, France.
Biochim Biophys Acta. 2008 Jan;1778(1):105-12. doi: 10.1016/j.bbamem.2007.09.017. Epub 2007 Sep 29.
A commonly-used method for analysing raft membrane domains is based on their resistance to extraction by non-ionic detergents at 4 degrees C. However, the selectivity of different detergents in defining raft membrane domains has been questioned. We have compared the lipid composition of detergent-resistant membranes (DRMs) obtained after Triton X-100 or Lubrol WX extraction in MDCK cells in order to understand the differential effect of these detergents on membranes and their selectivity in solubilizing or not proteins. Both Lubrol and Triton DRMs were enriched with cholesterol over the lysate, thus exhibiting characteristics consistent with the properties of membrane rafts. However, the two DRM fractions differed considerably in the ratio between lipids of the inner and outer membrane leaflets. Lubrol DRMs were especially enriched with phosphatidylethanolamine, including polyunsaturated species with long fatty acyl chains. Lubrol and Triton DRMs also differed in the amount of raft transmembrane proteins and raft proteins anchored to the cytoplasmic leaflet. Our results suggest that the inner side of rafts is enriched with phosphatidylethanolamine and cholesterol, and is more solubilized by Triton X-100 than by Lubrol WX.
一种常用的分析筏膜结构域的方法是基于它们在4摄氏度下对非离子型去污剂提取的抗性。然而，不同去污剂在定义筏膜结构域方面的选择性受到了质疑。我们比较了在MDCK细胞中用Triton X-100或Lubrol WX提取后获得的抗去污剂膜（DRM）的脂质组成，以了解这些去污剂对膜的不同影响及其在溶解或不溶解蛋白质方面的选择性。与裂解物相比，Lubrol和Triton DRM都富含胆固醇，因此表现出与膜筏特性一致的特征。然而，这两个DRM组分在内膜和外膜小叶脂质比例上有很大差异。Lubrol DRM特别富含磷脂酰乙醇胺，包括具有长脂肪酰链的多不饱和物种。Lubrol和Triton DRM在筏跨膜蛋白和锚定在细胞质小叶上的筏蛋白数量上也有所不同。我们的结果表明，筏的内侧富含磷脂酰乙醇胺和胆固醇，并且用Triton X-100比用Lubrol WX更易溶解。
