SEPARATION AND CHARACTERIZATION OF GLYCOPROTEIN AND GLYCOLIPID OLIGOSACCHARIDES

糖蛋白和糖脂低聚糖的分离和表征

• 批准号: 8363033
• 负责人: Parastoo Azadi
• 金额: $ 0.34万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8363033
• 关键词: Anions; Area; Biochemical; Biological Assay; Capillary Electrophoresis; Carbohydrates; Chromatography; Detection; Digestion; Educational process of instructing; Endoglycosidases; Exoglycosidases; Faculty; Funding; Glycoconjugates; Glycolipids; Glycoproteins; Grant; High Pressure Liquid Chromatography; Institutes; Laboratories; Lectin; Literature; Mass Spectrum Analysis; Monosaccharides; NMR Spectroscopy; National Center for Research Resources; Oligosaccharides; Polysaccharides; Principal Investigator; Protocols documentation; Research; Research Infrastructure; Resources; Site; Source; Structure; Techniques; Technology; Thin Layer Chromatography; United States National Institutes of Health; cost; experience; glycosylation; instrument; lectures; sugar

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Separation and Characterization of Glycoprotein and Glycolipid Oligosaccharides. This is a one-week course that covers basic techniques for the isolation and characterization of oligosaccharides, including colorimetric quantitation of sugars, chromatographic separation of oligosaccharide mixtures by HPLC, Metrohm peak high-pH anion-exchange chromatography for monosaccharide composition analysis, enzymatic release of oligosaccharides from a glycoconjugate, separation of glycolipids by thin-layer chromatography and detection by overlay assay, and endo- and exoglycosidase treatment of oligosaccharides. The use of lectin blotting techniques for characterization of oligosaccharide structural features are covered. Other topics included are conjugation of oligosaccharides with fluorescent tags, capillary electrophoresis and monosaccharide composition analysis. In addition, modules on using mass spectrometry and/or NMR spectroscopy for analysis of glycoconjugates are included (see descriptions below). Experience with basic biochemical techniques is a prerequisite for participation. In addition to CCRC faculty, other speakers were invited to help teach the course. Dr. Alfred Merrill from Georgia Institute of Technology gave a lecture on "Structural Analysis of Glycolipids by Mass Spectrometry". Additional Course Modules Mass Spectrometric (MS) Analysis of Glycoconjugates: The use of mass spectrometry for the characterization of glycoconjugates is discussed. Topics in this area include identifying occupied glycosylation sites by LC-MS and sequencing glycoprotein glycans with sequential exoglycosidase digestions followed by MS. Laboratory demonstrations are performed on these two topics. NMR of Carbohydrates: The NMR module consists of an introduction to the application of NMR spectroscopy to polysaccharides and glycoconjugates covering (i) essential features of NMR spectra, (ii) protocols used to solve primary structures, (iii) applications and examples from the current literature, and (iv) resources for information. Our four NMR instruments (300-, 500-, 600-, and 800-MHz) are used for demonstrations during this module.

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其他NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， NCRR赠款不直接向子项目或子项目工作人员提供资金。 糖蛋白和糖脂寡聚体的分离与表征。 这是一个为期一周的课程，涵盖了低聚糖分离和表征的基本技术，包括糖的比色定量，低聚糖混合物的HPLC色谱分离，用于单糖组成分析的瑞士万通峰高pH阴离子交换色谱法，从糖缀合物中酶促释放低聚糖，薄层色谱法分离糖脂和重叠测定法检测，以及寡糖的内切和外切糖苷酶处理。使用凝集素印迹技术表征寡糖的结构特征。其他主题包括低聚糖与荧光标记的结合，毛细管电泳和单糖组成分析。此外，还包括使用质谱法和/或NMR光谱法分析糖缀合物的模块（见下文描述）。具有基本生物化学技术的经验是参加的先决条件。除了CCRC教师，其他演讲者被邀请来帮助教授课程。来自格鲁吉亚理工学院的Alfred梅里尔博士作了题为“糖脂的质谱结构分析”的讲座。 附加课程模块 糖缀合物的质谱（MS）分析：讨论了糖缀合物表征的质谱法的使用。在这方面的主题包括确定占领的糖基化位点的LC-MS和测序糖蛋白聚糖与顺序外切糖苷酶digestion随后由MS。实验室演示进行这两个主题。 碳水化合物的NMR：NMR模块包括介绍NMR光谱在多糖和糖缀合物中的应用，涵盖（i）NMR光谱的基本特征，（ii）用于解决一级结构的协议，（iii）当前文献中的应用和示例，以及（iv）信息资源。我们的四台NMR仪器（300-，500-，600-和800-MHz）用于本模块的演示。