Bolard J, Legrand P, Heitz F, Cybulska B
Laboratoire de Physique et Chimie Biomoléculaires (U.A. C.N.R.S. 198), Université Pierre et Marie Curie, Paris, France.
Biochemistry. 1991 Jun 11;30(23):5707-15. doi: 10.1021/bi00237a011.
The inducement of K+ permeability through membranes by the polyene antibiotic amphotericin B (AmB) has been analyzed as a measure of the antibiotic activity. Dose-response curves have been obtained with cholesterol- and ergosterol-containing egg yolk phosphatidylcholine large unilamellar vesicles (LUVs), human erythrocytes, and Saccharomyces cerevisiae cells. Conductance changes induced by AmB in sterol-containing planar bilayer membranes have also been studied. AmB self-association in aqueous buffer was determined by circular dichroism (CD) as a function of the antibiotic concentration. Electronic absorption and CD spectra of AmB were recorded in the presence of LUVs. For given AmB concentrations, the extent of permeability inducement is dependent on the lipid concentration. On the other hand, for cholesterol-containing LUVs or erythrocytes, a critical AmB concentration had to be reached before any permeability is observed. Independent of lipid concentration, this concentration was directly related to antibiotic self-association in the aqueous buffer. The same observation was made for erythrocytes and nystatin. The AmB absorption and CD spectra were totally different for ergosterol- and cholesterol-containing LUVs. Formation of single channels by one-sided addition of AmB could be observed only in ergosterol-containing membranes. These data lead us to propose that the permeability pathways induced by amphotericin B or nystatin, in ergosterol- and in cholesterol-containing membranes, are of different natures. In the latter case the antibiotics are only active, by single-sided addition, in the self-associated form. These findings offer important clues for the design of less toxic derivatives of AmB: they should have a low degree of self-association in water.
多烯抗生素两性霉素B（AmB）诱导钾离子透过膜的作用已被分析，以此作为衡量抗生素活性的一种方法。已获得了含胆固醇和麦角固醇的蛋黄磷脂酰胆碱大单层囊泡（LUVs）、人红细胞和酿酒酵母细胞的剂量反应曲线。还研究了AmB在含固醇的平面双层膜中诱导的电导变化。通过圆二色性（CD）测定了AmB在水性缓冲液中的自缔合作用，该作用是抗生素浓度的函数。在LUVs存在的情况下记录了AmB的电子吸收光谱和CD光谱。对于给定的AmB浓度，通透性诱导程度取决于脂质浓度。另一方面，对于含胆固醇的LUVs或红细胞，在观察到任何通透性之前必须达到临界AmB浓度。与脂质浓度无关，该浓度与水性缓冲液中抗生素的自缔合直接相关。对红细胞和制霉菌素也有相同的观察结果。对于含麦角固醇和含胆固醇的LUVs，AmB的吸收光谱和CD光谱完全不同。仅在含麦角固醇的膜中可以观察到通过单侧添加AmB形成单通道。这些数据使我们提出，两性霉素B或制霉菌素在含麦角固醇和含胆固醇的膜中诱导的通透性途径具有不同的性质。在后一种情况下，通过单侧添加，抗生素仅以自缔合形式具有活性。这些发现为设计毒性较小的AmB衍生物提供了重要线索：它们在水中应具有低程度的自缔合。
