Selinsky B S, Zhou Z, Fojtik K G, Jones S R, Dollahon N R, Shinnar A E
Department of Chemistry, Villanova University, Villanova, PA 19085-1699, USA.
Biochim Biophys Acta. 1998 Mar 13;1370(2):218-34. doi: 10.1016/s0005-2736(97)00265-4.
The ability of the shark antimicrobial aminosterol squalamine to induce the leakage of polar fluorescent dyes from large unilamellar phospholipid vesicles (LUVs) has been measured. Micromolar squalamine causes leakage of carboxyfluorescein (CF) from vesicles prepared from the anionic phospholipids phosphatidylglycerol (PG), phosphatidylserine (PS), and cardiolipin. Binding analyses based on the leakage data show that squalamine has its highest affinity to phosphatidylglycerol membranes, followed by phosphatidylserine and cardiolipin membranes. Squalamine will also induce the leakage of CF from phosphatidylcholine (PC) LUVs at low phospholipid concentrations. At high phospholipid concentrations, the leakage of CF from PC LUVs deviates from a simple dose-response relationship, and it appears that some of the squalamine can no longer cause leakage. Fluorescent dye leakage generated by squalamine is graded, suggesting the formation of a discrete membrane pore rather than a generalized disruption of vesicular membranes. By using fluorescently labeled dextrans of different molecular weight, material with molecular weight </=4000 g/mol is released from vesicles by squalamine, but material with molecular weight >/=10,000 is retained. Negative stain electron microscopy of squalamine-treated LUVs shows that squalamine decreases the average vesicular size in a concentration-dependent manner. Squalamine decreases the size of vesicles containing anionic phospholipid at a lower squalamine/lipid molar ratio than pure PC LUVs. In a centrifugation assay, squalamine solubilizes phospholipid, but only at significantly higher squalamine/phospholipid ratios than required for either dye leakage or vesicle size reduction. Squalamine solubilizes PC at lower squalamine/phospholipid ratios than PG. We suggest that squalamine complexes with phospholipid to form a discrete structure within the bilayers of LUVs, resulting in the transient leakage of small encapsulated molecules. At higher squalamine/phospholipid ratios, these structures release from the bilayers and aggregate to form either new vesicles or squalamine/phospholipid mixed micelles.
鲨鱼抗菌氨基甾醇角鲨胺诱导极性荧光染料从大单层磷脂囊泡（LUVs）泄漏的能力已被测定。微摩尔浓度的角鲨胺会导致羧基荧光素（CF）从由阴离子磷脂磷脂酰甘油（PG）、磷脂酰丝氨酸（PS）和心磷脂制备的囊泡中泄漏。基于泄漏数据的结合分析表明，角鲨胺对磷脂酰甘油膜的亲和力最高，其次是磷脂酰丝氨酸膜和心磷脂膜。在低磷脂浓度下，角鲨胺也会诱导CF从磷脂酰胆碱（PC）LUVs中泄漏。在高磷脂浓度下，CF从PC LUVs的泄漏偏离简单的剂量反应关系，似乎一些角鲨胺不再能引起泄漏。角鲨胺产生的荧光染料泄漏是分级的，表明形成了离散的膜孔，而不是囊泡膜的普遍破坏。通过使用不同分子量的荧光标记葡聚糖，分子量≤4000 g/mol的物质会被角鲨胺从囊泡中释放出来，但分子量≥10,000的物质会被保留。对角鲨胺处理的LUVs进行负染电子显微镜观察表明，角鲨胺以浓度依赖的方式降低了平均囊泡大小。与纯PC LUVs相比，角鲨胺以更低的角鲨胺/脂质摩尔比降低了含有阴离子磷脂的囊泡大小。在离心测定中，角鲨胺可溶解磷脂，但所需的角鲨胺/磷脂比显著高于染料泄漏或囊泡大小减小所需的比例。与PG相比，角鲨胺以更低的角鲨胺/磷脂比溶解PC。我们认为，角鲨胺与磷脂结合，在LUVs的双层膜内形成离散结构，导致小的包封分子暂时泄漏。在较高的角鲨胺/磷脂比下，这些结构从双层膜中释放出来并聚集形成新的囊泡或角鲨胺/磷脂混合胶束。
