Growth and transformation of vesicles studied by ferritin labeling and cryotransmission electron microscopy
Growth and transformation of vesicles studied by ferritin labeling and cryotransmission electron microscopy
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DOI：
10.1021/jp001298i
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发表时间：
2001-02-08
影响因子：
3.3
通讯作者：
Luisi, PL
中科院分区：
文献类型：
--
作者：
Berclaz, N;Müller, M;Luisi, PL
对油酸/油酸盐和 POPC/油酸/油酸盐囊泡（POPC 代表 1-棕榈酰-2-油酰-sn-甘油-3-磷酸胆碱）的生长和分裂过程进行了研究。初步地，动力学比浊研究表明，少量的POPC囊泡（[POPC]/[油酸+油酸盐] = 1:25）的存在大大加速了油酸胶束自发形成囊泡的非常缓慢的过程，这表明在后一种情况下，囊泡形成的过程完全不同。然后通过冷冻透射电子显微镜 (cryo-TEM) 并使用包埋的蛋白质铁蛋白来研究预形成囊泡大小分布的变化，铁蛋白由于其致密的铁核心，可以通过冷冻 TEM 进行检测。通过这种方式，确定了“填充”（含铁蛋白）和“空”囊泡的尺寸分布。在预先形成的 POPC 囊泡中添加油酸盐导致含铁蛋白囊泡的尺寸分布向更大直径方向转变，同时囊泡数量也增加。大多数这些囊泡是空的、单层的，并且通常比预先形成的囊泡小。这表明新的囊泡是通过较大囊泡的裂变过程产生的，而较大囊泡是由预先形成的囊泡产生的。通过油酸酐水解原位产生的油酸和油酸盐分子（作为上清液添加到含有预先形成的含铁蛋白的油酸/油酸盐囊泡的水悬浮液中）导致囊泡尺寸分布变宽和空囊泡百分比增加。所有数据一起提供了添加新鲜表面活性剂后囊泡生长的证据，以及导致大量小囊泡的较大囊泡裂变过程的证据。
An investigation of the processes of growth and division of oleic acid/oleate and POPC/oleic acid/oleate vesicles (POPC stands for 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) is presented. Preliminarily, kinetic turbidimetric studies show that the very slow process of spontaneous formation of vesicles from oleote micelles is dramatically accelerated by the presence of a small amount of performed POPC vesicles ([POPC]/[oleic acid + oleate] = 1:25), suggesting in the latter case a quite different process of vesicle formation. The changes in the size distribution of preformed vesicles is then studied by cryotransmission electron microscopy (cryo-TEM) and by using an entrapped protein, ferritin, which, due to its dense iron core, can be detected by cryo-TEM. In this way, the size distributions of "filled" (ferritin-containing) and "empty" vesicles were determined. Addition of oleate to preformed POPC vesicles resulted in a shift of the size distribution of the ferritin-containing vesicles toward larger diameters, together with an increase in the number of vesicles. Most of these vesicles were empty, unilamellar, and often smaller than the preformed ones. This suggests the generation of new vesicles through a fission process of larger vesicles, which arise from the preformed vesicles. Oleic acid and oleate molecules produced in situ through the hydrolysis of oleic anhydride (added as a supernatant to an aqueous suspension containing preformed ferritin-containing oleic acid/oleate vesicles) led to a broadening of the vesicle size distribution and an increase of the percentage of empty vesicles. All of the data taken together give evidence of the growth of vesicles upon addition of fresh surfactant, as well as evidence of fission processes of larger vescicles that lead to a large number of small vesicles.