How to extract selectively the lo-phase domains from large unilamellar vesicles with Triton X-100?
How to extract selectively the lo-phase domains from large unilamellar vesicles with Triton X-100?
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DOI：
10.1016/j.colsurfa.2005.12.044
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发表时间：
2006-07-20
影响因子：
5.2
通讯作者：
Puff, Nicolas
中科院分区：
文献类型：
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作者：
Coste, Virginie;Breton, Michelyne;Puff, Nicolas
诱导的结构过渡阶段的结果，在4度C的Triton X-100与大的单层囊泡（LUVs）的相互作用进行了研究，通过蔗糖浮选程序类似于用于分离生物洗涤剂抗性膜（DRM）。基于40-35-5%蔗糖密度梯度的每个级分的[1 α，2 α（n）-H-3]-胆固醇含量，测定离心后脂质结构的漂浮。我们测量了大单层囊泡的不同脂质组成和不同有效去污剂与脂质比的Triton X-100不溶性漂浮部分（TIFFS）的量。在4 ℃下，对于双组分脂质膜（PC/SM 2：1，mol/mol），50的有效去污剂与脂质比是完成膜溶解所必需的。当液体有序和液体无序相域共存于囊泡膜（PC/SM/Chol 53：27：20，mol/mol），完全增溶发生在更高的有效去污剂脂质比。这与液体有序相对洗涤剂萃取的较高抗性一致。然而，在异质（l（o）/l（d）相）囊泡的情况下，并为一系列有效的洗涤剂脂质比例促进不完全溶解，我们检测到TIFF中的中间密度结构，不存在双组分脂质膜（PC/SM 2：1，mol/mol）。我们解释了这些结果与我们小组最近的研究结果，并提出了一种机制，异质性大单层囊泡溶解。我们表明，对于脂质双层表现出l（o）/l（d）相共存，一个特定的有效的洗涤剂与脂质的比例，允许预先存在的紧密堆积的有序结构域的隔离，可以发现，但在任何情况下，一定量的洗涤剂是在浮动部分膜。(c)2006 Elsevier B. V.保留所有权利。
The structural transition stages induced as a result of interaction at 4 degrees C of the Triton X-100 with large unilamellar vesicles (LUVs) were studied by means of a sucrose flotation procedure similar to that used to isolate biological detergent resistant membranes (DRMs). Flotation of lipid structures after centrifugation was determined on the basis of the [1 alpha, 2 alpha (n)-H-3]-cholesterol content of each fraction of a 40-35-5% sucrose density gradient. We measured the amount of Triton X-100 insoluble floating fractions (TIFFs) for different lipid compositions of large unilamellar vesicles and different effective detergent to lipid ratios. At 4 degrees C and for two-component lipid membrane (PC/SM 2:1,mol/mol), an effective detergent to lipid ratio of 50 is necessary to complete membrane solubilization. When liquid-ordered and liquid-disordered phase domains coexist in the vesicle membrane (PC/SM/Chol 53:27:20, mol/mol), complete solubilization occurs at higher effective detergent to lipid ratio. This is consistent with a higher resistance of the liquid-ordered phase to detergent extraction. Nevertheless, in the case of heterogeneous (l(o)/l(d) phase) vesicles, and for a range of effective detergent to lipid ratios promoting incomplete solubilization, we detected in TIFF intermediate density structures which did not exist for two-component lipid membranes (PC/SM 2:1, mol/mol). We interpreted these results in relation with recent findings of our group and propose a mechanism for heterogeneous large unilamellar vesicle solubilization. We show that for lipid bilayers exhibiting l(o)/l(d) phase co-existence, a specific effective detergent to lipid ratio allowing the isolation of pre-existing tightly packed ordered domains can be found, but, in any case, certain amount of the detergent is presented in floating fraction membranes. (c) 2006 Elsevier B.V. All rights reserved.