Liu Yu, Drickamer Kurt, Taylor Maureen E
Department of Life Sciences, Sir Ernst Chain Building, Imperial College London, Exhibition Road, London SW7 2AZ, United Kingdom.
Glycobiology. 2024 Dec 10;34(12). doi: 10.1093/glycob/cwae083.
The sugar-binding receptor mincle stimulates macrophages when it encounters surface glycans on pathogens, such as trehalose dimycolate glycolipid in the outer membrane of mycobacteria. Binding of oligosaccharide ligands to the extracellular C-type carbohydrate-recognition domain (CRD) in mincle initiates intracellular signaling through the common Fc receptor γ (FcRγ) adapter molecule associated with mincle. One potential mechanism for initiation of signaling involves clustering of receptors, so it is important to understand the oligomeric state of mincle. Affinity purification of mincle from transfected mammalian cells has been used to show that mincle exists as a pre-formed, disulfide-linked dimer. Deletion of cysteine residues and chemical crosslinking further demonstrate that the dimers of mincle are stabilized by a disulfide bond between cysteine residues in the neck sequence that links the CRD to the membrane. In contrast, cysteine residues in the transmembrane region of mincle are not required for dimer formation or association with FcRγ. A protocol has been developed for efficient production of a disulfide-linked extracellular domain fragment of mincle in a bacterial expression system by appending synthetic dimerization domains to guide dimer formation in the absence of the membrane anchor.
糖结合受体小胶质细胞诱导物（Mincle）在遇到病原体表面聚糖时会刺激巨噬细胞，比如分枝杆菌外膜中的海藻糖二霉菌酸糖脂。寡糖配体与Mincle细胞外C型碳水化合物识别结构域（CRD）结合，通过与Mincle相关的共同Fc受体γ（FcRγ）衔接分子启动细胞内信号传导。信号传导起始的一种潜在机制涉及受体聚集，因此了解Mincle的寡聚状态很重要。从转染的哺乳动物细胞中亲和纯化Mincle已用于表明Mincle以预先形成的、二硫键连接的二聚体形式存在。半胱氨酸残基的缺失和化学交联进一步证明，Mincle的二聚体通过连接CRD与膜的颈部序列中的半胱氨酸残基之间的二硫键得以稳定。相比之下，Mincle跨膜区域中的半胱氨酸残基对于二聚体形成或与FcRγ的结合并非必需。通过附加合成二聚化结构域以在没有膜锚定的情况下引导二聚体形成，已开发出一种在细菌表达系统中高效生产Mincle二硫键连接的细胞外结构域片段的方案。
