de Bony J, Tocanne J F
Eur J Biochem. 1984 Sep 3;143(2):373-9. doi: 10.1111/j.1432-1033.1984.tb08383.x.
It is shown that photo-induced cross-linking reaction between anthracene-labelled phospholipids can be used for studying, at a molecular level, their lateral distribution in bilayer structures. A simple and versatile method is proposed. It is based on the property of anthracene to form covalently bound dimers upon irradiation in the near ultraviolet (360 nm) and on the possibility of separating the lipid photo-dimers from the lipid monomers by thin-layer chromatography. Identification of the photo-dimers is easily achieved since, upon illumination at a shorter wavelength (250-280 nm), they partially dissociate to the native monomer molecules. The feasibility of the method was tested by checking the effects of cations (sodium, calcium) on the homogeneity of 1/1 mixtures of anthracene-phosphatidylcholine, i.e. 1-acyl-2-[9-(2-anthryl)-nonanoyl]-sn-glycero-3-phosphocholin e (Anthr-PC) with anthracene-phosphatidic acid (Anthr-PA) and with anthracene-phosphatidylglycerol (Anthr-PG) in the form of liposomes. These lipids were anthracene-labelled by acylation of their glycerol backbone at the sn-2 position with the synthetic 9-(2-anthryl)-nonanoic acid. Data presented indicate a good miscibility of these lipids in the presence of sodium. For each lipid mixture, the lipid heterodimers were clearly identified and, quantitatively, they dominated the lipid homodimers, as expected for a regular distribution of the lipids in the 1/1 mixture. Addition of calcium ions to the lipid suspensions did not alter the miscibility properties of Anthr-PC and Anthr-PG. In contrast, calcium triggered a clear-cut phase separation in the Anthr-PC/Anthr-PA mixture as, in this case, only traces of the heterodimer form of the lipids remained observable on the chromatogram. The three anthracene-phospholipids, pure or mixed together, exhibit a clear-cut gel-to-liquid phase transition which was detectable by fluorescence intensity measurements. The analysis of the corresponding phase-transition temperatures confirms, at a 'macroscopic' level, the effects of sodium and calcium on the mixing properties of the anthracene phospholipids which were revealed at a 'microscopic' level by the dimerization procedure.
结果表明，蒽标记的磷脂之间的光诱导交联反应可用于在分子水平上研究它们在双层结构中的横向分布。本文提出了一种简单且通用的方法。该方法基于蒽在近紫外光（360nm）照射下形成共价结合二聚体的性质，以及通过薄层色谱法将脂质光二聚体与脂质单体分离的可能性。由于光二聚体在较短波长（250 - 280nm）照射下会部分解离为天然单体分子，因此很容易对光二聚体进行鉴定。通过检查阳离子（钠、钙）对蒽 - 磷脂酰胆碱（即1 - 酰基 - 2 - [9 - （2 - 蒽基） - 壬酰基] - sn - 甘油 - 3 - 磷酸胆碱（Anthr - PC））与蒽 - 磷脂酸（Anthr - PA）和蒽 - 磷脂酰甘油（Anthr - PG）以脂质体形式存在的1/1混合物均匀性的影响，测试了该方法的可行性。这些脂质通过在其甘油主链的sn - 2位置用合成的9 - （2 - 蒽基） - 壬酸进行酰化反应而被蒽标记。所呈现的数据表明这些脂质在钠存在下具有良好的互溶性。对于每种脂质混合物，脂质异二聚体都能被清晰鉴定，并且从数量上看，它们在脂质同二聚体中占主导地位，这与脂质在1/1混合物中的规则分布预期相符。向脂质悬浮液中添加钙离子不会改变Anthr - PC和Anthr - PG的互溶性质。相反，钙会引发Anthr - PC/Anthr - PA混合物中明显的相分离，因为在这种情况下，色谱图上只能观察到痕量的脂质异二聚体形式。这三种蒽 - 磷脂，无论是纯的还是混合在一起，都表现出明显的凝胶 - 液相转变，这可以通过荧光强度测量检测到。对相应相变温度的分析在“宏观”水平上证实了钠和钙对蒽磷脂混合性质所产生的影响，而这些影响在“微观”水平上已通过二聚化程序揭示出来。
