Abstract:
Natamycin is a polyene macrolide, widely employed to treat fungal keratitis and other yeast infections as well as to protect food products against fungal molds. In contrast to other polyene macrolides, such as nystatin or amphotericin B, natamycin does not form pores in yeast membranes, and its mode of action is not well understood. Here, we have employed a variety of spectroscopic methods, computational modeling, and membrane reconstitution to study the molecular interactions of natamycin underlying its antifungal activity. We find that natamycin forms aggregates in an aqueous solution with strongly altered optical properties compared to monomeric natamycin. Interaction of natamycin with model membranes results in a concentration-dependent fluorescence increase which is more pronounced for ergosterol- compared to cholesterol-containing membranes up to 20 mol% sterol. Evidence for formation of specific ergosterol-natamycin complexes in the bilayer is provided. Using nuclear magnetic resonance (NMR) and electron spin resonance (ESR) spectroscopy, we find that natamycin sequesters sterols, thereby interfering with their well-known ability to order acyl chains in lipid bilayers. This effect is more pronounced for membranes containing the sterol of fungi, ergosterol, compared to those containing mammalian cholesterol. Natamycin interferes with ergosterol-dependent transport of lysine by the yeast transporter Lyp1, which we propose to be due to the sequestering of ergosterol, a mechanism that also affects other plasma membrane proteins. Our results provide a mechanistic explanation for the selective antifungal activity of natamycin, which can set the stage for rational design of novel polyenes in the future.
摘要:
纳他霉素是一种多烯大环内酯,广泛用于治疗真菌性角膜炎和其他酵母菌感染,以及保护食品免受真菌霉菌。与其他多烯大环内酯相比,如制霉菌素或两性霉素B,纳他霉素不会在酵母膜中形成孔,它的作用方式还没有得到很好的理解。这里,我们采用了多种光谱方法,计算建模,和膜重建以研究纳他霉素抗真菌活性的分子相互作用。我们发现游霉素在水溶液中形成聚集体,与单体游霉素相比,其光学特性发生了强烈变化。游霉素与模型膜的相互作用导致浓度依赖性荧光增加,与含胆固醇的膜高达20mol%的固醇相比,麦角甾醇的浓度依赖性荧光增加更为明显。提供了在双层中形成特定麦角甾醇-游霉素复合物的证据。使用核磁共振(NMR)和电子自旋共振(ESR)光谱,我们发现纳他霉素螯合甾醇,从而干扰它们众所周知的在脂质双层中排序酰基链的能力。这种效果对于含有真菌甾醇的膜更明显,麦角固醇,与含有哺乳动物胆固醇的动物相比。纳他霉素通过酵母转运蛋白Lyp1干扰麦角甾醇依赖性赖氨酸的转运,我们认为这是由于麦角甾醇的螯合所致,一种也影响其他质膜蛋白质的机制。我们的结果为那他霉素的选择性抗真菌活性提供了机制解释,这可以为未来新型多烯的合理设计奠定基础。