Reconstitution of membranes with individual paramyxovirus glycoproteins and phospholipid in cholate solution.
Reconstitution of membranes with individual paramyxovirus glycoproteins and phospholipid in cholate solution.
复制标题
用胆酸盐溶液中的单个副粘病毒糖蛋白和磷脂重建膜。
DOI：
10.1016/0042-6822(79)90502-6
复制
发表时间：
1979
期刊：
影响因子：
3.7
通讯作者：
P. Choppin
中科院分区：
文献类型：
--
作者：
M. Hsu;A. Scheid;P. Choppin
发展了一种用仙台病毒单个糖蛋白(HN和F)和磷脂酰胆碱重建生物活性膜的方法。在Triton X-100存在下分离糖蛋白，并通过沉淀到含有2%胆酸盐的蔗糖梯度中转移到胆酸溶液中。然后用HN或F和磷脂酰胆碱通过透析去除胆酸盐来重建膜。用放射性标记洗涤剂的实验表明，Triton X-100通过沉积到胆酸盐中的去除和通过透析去除胆酸盐基本上是完全的。随着F蛋白的形成，根据初始混合物中脂肪和蛋白质的比例，形成了直径400-800？的囊泡和长度600-800？的细丝。这两个结构都覆盖着糖蛋白尖峰。对于HN蛋白，只形成小泡，表面刺突的密度取决于初始脂蛋白比。HN蛋白重组膜具有血凝活性和神经氨酸酶活性。当提供了一种将F蛋白-脂质复合体附着到细胞上的机制时，即通过添加小麦胚芽凝集素，含有F蛋白的重组颗粒显示出溶血活性。这些结果证实了F蛋白在膜融合中的作用，并表明对F蛋白活性的要求是先前证明的F蛋白的蛋白分解过程，F蛋白插入到脂质中，以及存在附着机制。
A method has been developed for reconstitution of biologically active membranes with individual Sendai virus glycoproteins (HN and F) and phosphatidylcholine. The glycoproteins were isolated in the presence of Triton X-100 and transferred into cholate solution by sedimentation into a sucrose gradient containing 2% cholate. Membranes were then reconstituted with HN or F and phosphatidylcholine by removal of cholate by dialysis. Experiments with radioactively labeled detergents showed that the removal of Triton X-100 by sedimentation into cholate and the removal of cholate by dialysis were essentially complete. With the F protein, vesicles 400–800 Å in diameter and filaments 600–800 Å in length were formed, depending on the proportions of lipid and protein in the initial mixture. Both structures were covered with glycoprotein spikes. With the HN protein, only vesicles were formed, and the densities of the spikes on the surface was dependent on the initial lipid to protein ratio. Membranes reconstituted from HN protein exhibited hemagglutinating and neuraminidase activities. Reconstituted particles containing the F protein exhibited hemolytic activity when a mechanism was provided to attach the F protein-lipid complex to the cell, i.e., by the addition of wheat germ agglutinin. These results have confirmed the role of the F protein in membrane fusion and have shown that the requirements for F protein activity are the previously demonstrated proteolytic processing of F, the insertion of the F protein into lipid, and the presence of an attachment mechanism.