Chemical Tools for the Investigation and Manipulation of Protein Glycosylation

摘要-“研究和操纵蛋白质糖基化的化学工具” 该提案的总体目标是开发化学工具，使人们能够轻松、稳健地 识别和抑制特定细胞中的糖基化，糖基化介导的和细胞- 类型特异性相互作用，以及碳水化合物生物合成途径的监测和操纵。的 糖基化是蛋白质翻译后修饰的最常见形式之一。 修饰并与各种过程相关，包括蛋白质稳定性，大分子 相互作用和细胞信号传导。不幸的是，目前可用于查询这些功能的工具 这使得糖基化的研究仅限于专家实验室。我们计划通过以下方式来满足这一未满足的需求： 碳水化合物化学，光化学和化学生物学三个具体目标。在目标1中，我们 建立在我们的糖基化探针和抑制剂的发展，重点是使用化学遗传学 创建工具的方法，以细胞特异性方式识别和干扰糖基化。在目标2中，我们将 利用糖基化的化学酶修饰的优势， 在活细胞上，重点是识别由聚糖介导的生物相互作用，以及 绘制细胞相互作用。最后，在目标3中，我们将创建新的基于活动的探针，用于测量和 抑制负责单糖生物合成的关键酶。在这些结束时， 独立的目标，我们将产生新的强大的工具，将有直接的影响类型 关于糖基化在人类健康和疾病中的作用，科学家们可以提出许多问题。

ABSTRACT - “Chemical Tools for the Investigation and Manipulation of Protein Glycosylation” The broad goal of this proposal is the development of chemical tools that will enable the facile and robust identification and inhibition of glycosylation in specific cells, the mapping of glycosylation-mediated and cell- type specific interactions, and monitoring and manipulation of carbohydrate biosynthetic pathways. The addition of carbohydrates to proteins, or glycosylation, is one of the most common forms of posttranslational modifications and is associated with various processes, including protein stability, macromolecular interactions, and cellular signaling. Unfortunately, the currently available tools for interrogating these functions fall short, which limits the study of glycosylation to expert labs. We plan to tackle this unmet need using carbohydrate chemistry, photo-chemistry, and chemical biology in three specific aims. In Aim 1, we will build on our development of glycosylation probes and inhibitors, with a focus on using chemical genetic approaches to create tools to identify and perturb glycosylation in a cell-specific fashion. In Aim 2, we will leverage the advantages of chemoenzymatic modification of glycosylation to install specific photocrosslinkers onto living cells, with a focus on identifying biological interactions that are mediated by glycans, as well as mapping cell interactions. Finally, in Aim 3, we will create novel activity-based probes for measuring and inhibiting critical enzymes responsible for monosaccharide biosynthesis. At the conclusion of these independent aims, we will have generated new powerful tools that will have an immediate impact on the types of questions scientists can ask about glycosylation in human health and disease.