Solid-state NMR Reveals the Carbon-based Molecular Architecture of Cryptococcus neoformans Fungal Eumelanins in the Cell Wall
Solid-state NMR Reveals the Carbon-based Molecular Architecture of Cryptococcus neoformans Fungal Eumelanins in the Cell Wall
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DOI：
10.1074/jbc.m114.618389
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发表时间：
2015-05-29
影响因子：
4.8
通讯作者：
Stark, Ruth E.
中科院分区：
文献类型：
--
作者：
Chatterjee, Subhasish;Prados-Rosales, Rafael;Stark, Ruth E.
黑色素对电离辐射和自由基都有保护作用，并具有潜在的土壤修复能力。致病性新隐球菌真菌产生的真黑素是使真菌细胞抵抗宿主防御和某些抗真菌药物的毒力因子。由于它们的不溶性和无定形特性，无论是色素结合框架还是潜在的黑色素化的细胞相互作用，都没有得到全面的分子尺度研究。本研究利用C. neoformans对外源性儿茶酚胺前体黑色素化的需求，产生同位素富集的色素“鬼”，并应用2D C-13-C-13相关固态NMR揭示了真菌细胞中完整天然真黑素组装体的碳基结构。我们证明了固体新生C.黑色素鬼的脂肪族部分包括来自多糖和/或几丁质的细胞壁成分，这些成分与脂质膜成分近端相关。在成熟的芳香真菌色素发育之前，这些脂肪族部分形成了一个耐化学腐蚀的框架，可以作为黑色素合成的支架。基于吲哚的核心芳香族部分显示出与所提出的由堆叠平面组件组成的黑色素结构一致的互连，这些结构在空间上与脂肪族支架相关。颜料的吡咯族碳通过糖苷或甘油官能团与脂肪族框架共价结合。这些发现证实了色素组装的结构随着时间的推移而变化，并提供了关于黑色素在真菌细胞壁中组装机制的第一个生物物理信息，提供了重要的见解，可以推进生物启发导电纳米材料和新疗法的设计。
Melanin pigments protect against both ionizing radiation and free radicals and have potential soil remediation capabilities. Eumelanins produced by pathogenic Cryptococcus neoformans fungi are virulence factors that render the fungal cells resistant to host defenses and certain antifungal drugs. Because of their insoluble and amorphous characteristics, neither the pigment bonding framework nor the cellular interactions underlying melanization of C. neoformans have yielded to comprehensive molecular-scale investigation. This study used the C. neoformans requirement of exogenous obligatory catecholamine precursors for melanization to produce isotopically enriched pigment "ghosts" and applied 2D C-13-C-13 correlation solid-state NMR to reveal the carbon-based architecture of intact natural eumelanin assemblies in fungal cells. We demonstrated that the aliphatic moieties of solid C. neoformans melanin ghosts include cell-wall components derived from polysaccharides and/or chitin that are associated proximally with lipid membrane constituents. Prior to development of the mature aromatic fungal pigment, these aliphatic moieties form a chemically resistant framework that could serve as the scaffold for melanin synthesis. The indole-based core aromatic moieties show interconnections that are consistent with proposed melanin structures consisting of stacked planar assemblies, which are associated spatially with the aliphatic scaffold. The pyrrole aromatic carbons of the pigments bind covalently to the aliphatic framework via glycoside or glyceride functional groups. These findings establish that the structure of the pigment assembly changes with time and provide the first biophysical information on the mechanism by which melanin is assembled in the fungal cell wall, offering vital insights that can advance the design of bio-inspired conductive nanomaterials and novel therapeutics.