Chemical tools for studying fucosylated glycans

描述（由申请人提供）：浓缩聚糖广泛分布于真核生物和某些细菌中。在哺乳动物细胞表面，它们介导多种生理和病理过程，包括血管生成、受精、胚胎发生、炎症和肿瘤转移。在致病菌和寄生虫中，聚焦剂调节宿主组织的粘附和定植，并调节宿主的免疫反应。尽管聚焦聚糖具有明显的重要性，但由于其结构的复杂性和异质性，描述其功能的分子基础受到严重阻碍。目前，还没有一种简便、经济的化学方法来合成这些聚糖及其结构相关衍生物。该项目的长期目标是开发新的方法来制备结构明确的聚焦蛋白及其衍生物，并制造新的聚糖阵列平台，以全面探索聚焦蛋白与蛋白质的相互作用。在第一个授权期内，我们将侧重于三个具体目标。首先，我们将开发一种通用的化学酶策略，用于制备规模合成通用聚焦基供体，胍- 52-二磷酸-2- l -聚焦（GDP-聚焦），以及化学上定义的聚焦物及其结构相关衍生物。我们将利用聚焦激活和转移酶从细菌来源合成聚焦化聚糖和糖缀合物的功能研究。利用Aim 1中生成的focuside文库，我们将制备磷脂偶联糖树状大分子文库，并以微阵列形式将其整合到支持的脂质双层膜中（Aim 2）。平面脂质双分子层中聚糖的流体性质，加上多价显示，与当今大多数阵列中发现的常规固定化单体聚糖相比，更好地反映了自然界中发现的聚糖的呈现。结合糖树状聚合物微阵列技术，获得结构明确的聚焦蛋白提供了一种强大、快速的方法来分析聚焦蛋白与蛋白质的相互作用，并确定有助于结合的关键结构特征。在Aim 3中，我们将使用该技术识别非天然Lewis X衍生物，并增强DC-SIGN（树突状细胞特异性icam -3捕获非整合素）的亲和力，DC-SIGN是一种重要的内吞受体，介导抗原呈递。从本研究中鉴定出的具有增强DC-SIGN亲切度的聚糖配体将作为向树突状细胞运送货物的靶向元件在体外进行测试。

DESCRIPTION (provided by applicant): Fucosylated glycans are widely distributed throughout eukaryotes and certain bacteria. On the surface of mammalian cells, they mediate a variety of physiological and pathological processes, including angiogenesis, fertilization, embryogenesis, inflammation, and tumor metastasis. In pathogenic bacteria and parasites, fucosides regulate adhesion and colonization of host tissues and modulate the host immune response. Despite the obvious importance of fucosylated glycans, delineating the molecular basis of their function is severely hampered by their structural complexity and heterogeneity. Currently, there is no facile and cost-effective chemistry for synthesizing these glycans and their structurally related derivatives. The long term goal of this project is to develop new methods for the preparation of structurally defined fucosides and their derivatives and to fabricate new glycan array platforms for the comprehensive exploration of fucoside- protein interactions. In the first granting period, we will focus on three specific aims. First, we will develop a general chemoenzymatic strategy for preparative-scale synthesis of the universal fucosyl donor, guanidine 52- diphosphate-2-L-fucose (GDP-fucose), as well as chemically defined fucosides and their structurally related derivatives. We will harness fucosyl activation and transfer enzymes from bacterial sources to synthesize fucosylated glycans and glycoconjugates for their functional studies. Using the fucoside libraries generated in Aim 1, we will prepare a library of phospholipid-conjugated glycodendrimers and incorporate them into supported lipid bilayer membranes in a microarray format (Aim 2). The fluid nature of glycans in the planar lipid bilayer, coupled with the multivalent display, better mirrors the presentation of glycans found in nature as compared to the conventional immobilized monomeric glycans found in most arrays today. Access to structurally defined fucosides combined with the glycodendrimer microarray technology provides a powerful, rapid means to profile fucoside-protein interactions and to identify key structural features contributing to binding. In Aim 3, we will use this technology to identify unnatural Lewis X derivatives with enhanced avidity for DC-SIGN (dendritic cell-specific ICAM-3-grabbing nonintegrin), an important endocytic receptor mediating antigen presentation. The glycan ligands with enhanced DC-SIGN avidity identified from this study will be tested in vitro as targeting elements for delivering cargos to dendritic cells. PUBLIC HEALTH RELEVANCE: Fucosylated glycans are found on cell surfaces, where they play key roles in cell-cell interactions involved in normal biological processes and also in human disease. The goal of this research is to develop chemical tools for studying the biological functions of fucosylated glycans and glycoconjugates. These tools will improve our understanding of how these important glycans contribute to diseases such as cancer and inflammation.