Rapid Sequencing of Sulfated Glycans By CE-MS/MS

项目摘要/摘要 硫酸化糖胺聚糖(GAG)碳水化合物是结构最多样化的糖胺类化合物之一 生物分子及其生物结构-功能关系的全面了解还有待于 才能实现。与DNA/RNA和蛋白质等其他生物分子不同的是，这些生物分子是基于 模板，Gag生物合成是一系列生物合成酶累积作用的结果，这些酶 产生动态的、多分散的混合物。这种混合物的组成取决于以下因素 生物体年龄，发育或疾病状态，以及起源的组织。尽管这种多样性令人望而生畏 在分析挑战方面，该领域已取得重大进展，通过尝试分离和 表征从完整的多糖到酶法制备的寡糖和 二糖。最广泛的方法是通过分离(高效液相色谱， UHPLC、HILIC、CE)和检测(UV、荧光、质谱仪)。域可以是 以结构-功能研究为特点，通过将这些技术组合成连接的方法(例如LC- Ms)。尽管双糖分析是GAG分析中应用最广泛的方法，但它不能 在没有互补酶学的情况下，在参与蛋白质结合的GAG中提供结构基序 和领域建模。我们建议为GAG结构开发一个高吞吐量的CE-MS/MS平台 描述将使糖科学界能够广泛参与的方法 用于GAG寡糖分离的CE，一种新的CE-MS接口技术和稳健的串联质量 光谱分析接近了。

Project Summary/Abstract Sulfated glycosaminoglycan (GAG) carbohydrates represent one of the most structurally diverse groups of biomolecules and a comprehensive understanding of their biological structure-function relationships has yet to be achieved. Unlike other biomolecules such as DNA/RNA and proteins that are synthesized based upon a template, GAG biosynthesis is the result of the cumulative actions of a series of biosynthetic enzymes that produce a dynamic, polydisperse mixture. The composition of this mixture is dependent on factors such as organism age, developmental or disease state, and tissue of origin. Although this diversity presents a daunting analytical challenge, significant progress has been made in the field through attempts to isolate and characterize GAGs ranging from intact polysaccharides to enzymatically prepared oligosaccharides and disaccharides. The most widespread approach is to profile GAG disaccharides through separation (HPLC, UHPLC, HILIC, CE) and detection (UV, fluorescence, mass spectrometry (MS)). Domains can be characterized for structure-function studies by combining these techniques into hyphenated methods (e.g. LC- MS). Even though disaccharide analysis is the most widely utilized method for GAG analysis, it is unable to provide the structural motifs in GAGs that participate in protein binding without complementary enzymology and domain modeling. We propose to develop a high throughput CE-MS/MS platform for GAG structural characterization that will enable widespread engagement from the glycoscience community by incorporating CE for GAG oligosaccharide separation, a novel CE-MS interface technology, and robust tandem mass spectrometry approaches.