Characterization of glycan isomers by trapped ion mobility spectrometry-electron activated dissociation tandem mass spectrometry

捕获离子迁移谱-电子激活解离串联质谱法表征聚糖异构体

• 批准号: 9336322
• 负责人: Cheng Lin
• 金额: $ 31.91万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9336322
• 关键词: Affinity; Anabolism; Binding; Biological; Biological Process; Biopolymers; Capillary Electrophoresis; Cell surface; Cells; Common Core; Complement; Complex; Complex Mixtures; Computer software; Couples; Coupling; Databases; Deposition; Development; Devices; Dimensions; Disease; Dissociation; Electron Transport; Electrons; Endoplasmic Reticulum; Event; Fourier transform ion cyclotron resonance; Fractionation; Gases; Glycoproteins; Glycosaminoglycans; Golgi Apparatus; Health; Ions; Isomerism; Laboratories; Libraries; Liquid Chromatography; Mass Spectrum Analysis; Methods; Modification; Molecular Conformation; Nature; Neoplasm Metastasis; Oligonucleotides; Pattern; Performance; Phase; Play; Polysaccharides; Proteins; Resolution; Role; Sampling; Spectrometry; Stereoisomer; Structure; System; Techniques; Technology; Time; Tissues; analytical method; base; cell type; glycosylation; improved; improved mobility; instrument; interest; ion mobility; ionization; operation; pathogen; protein folding; tandem mass spectrometry; two-dimensional

Project summary/abstract Glycosylation plays vital roles in many cellular events, including protein folding, pathogen recognition, and cancer metastasis. The structural complexity and diversity of glycans parallel their diverse functions. Whereas the primary structures of linear biopolymers, such as proteins and oligonucleotides, are uniquely defined by their one-dimensional sequence, full structural characterization of a glycan requires determination of its two- dimensional topology, linkage and stereochemical configurations. Further analytical challenges arise from the non-template-driven nature of glycan biosynthesis, resulting in glycomes comprising a repertoire of closely- related structures, many of which structural isomers. Recently, a number of electron activated dissociation (ExD) methods have been developed in mass spectrometry laboratories for glycan analysis. Electron capture dissociation (ECD), electron transfer dissociation (ETD), and electronic excitation dissociation (EED) can yield rich structurally informative fragment ions for glycans analyzed in the positive ionization mode. In the negative ionization mode, electron detachment dissociation (EDD) and negative ETD (NETD) are powerful fragmentation methods for sequencing of acidic glycosaminoglycans. Meanwhile, ion mobility spectrometry (IMS) has been applied to separation of glycans. As a post-ionization, gas-phase separation method, IMS complements solution-phase separation methods such as capillary electrophoresis (CE) and liquid chromatography (LC), and can achieve isomer resolution based on differences in their gas-phase conformations. However, conventional drift-time IMS separation occurs on too short a time-scale to be compatible with the slower ExD analysis methods. A new IMS technique, termed trapped ion mobility spectrometry (TIMS), was recently introduced by Bruker Daltonics. We have demonstrated successful coupling of TIMS to high-performance Fourier-transform ion cyclotron resonance (FTICR) MS instrument for separation and identification of glycan linkage isomers. Here, we propose to modify the TIMS device and its control software, for improved mobility resolution, increased m/z operating range, and better integration with ExD-FTICR MS/MS analysis. We will then utilize the improved TIMS-ExD method for detailed structural characterization of glycans. We will also use TIMS-ExD MS/MS in conjunction with off-line LC fractionation to produce a library that contains identified glycan structures with their collision cross section values. This library will be made available to public. The initial development will be carried out on the FTICR MS platform, as it offers superior mass accuracy and resolving power, as well as the best ExD performance. The technology we develop here can later be transferred to other, more affordable MS instruments, following the development of alternative ECD cells to bring the ExD capability to non-ICR instruments.

项目概要/摘要 糖基化在许多细胞事件中起着至关重要的作用，包括蛋白质折叠、病原体识别和 癌症转移。聚糖的结构复杂性和多样性与其多样化的功能相平行。然而 线性生物聚合物（例如蛋白质和寡核苷酸）的一级结构由以下唯一定义 它们的一维序列，聚糖的完整结构表征需要确定其二维 维度拓扑、连接和立体化学构型。进一步的分析挑战来自 聚糖生物合成的非模板驱动性质，导致糖组包含一系列密切相关的 相关结构，其中许多是结构异构体。 近年来，大量电子激活解离（ExD）方法被开发出来。 用于聚糖分析的光谱实验室。电子捕获解离 (ECD)、电子转移 解离（ETD）和电子激发解离（EED）可以产生丰富的结构信息片段 在正电离模式下分析聚糖的离子。在负电离模式下，电子脱离 解离 (EDD) 和负 ETD (NETD) 是用于酸性测序的强大裂解方法 糖胺聚糖。同时，离子迁移谱（IMS）已应用于聚糖的分离。作为 IMS 是一种后电离气相分离方法，补充了溶液相分离方法，例如 如毛细管电泳（CE）和液相色谱（LC），并且可以基于 其气相构象的差异。然而，传统的漂移时间 IMS 分离也发生在 缩短时间尺度以与较慢的 ExD 分析方法兼容。一种新的 IMS 技术，称为 Bruker Daltonics 最近推出了俘获离子迁移谱 (TIMS)。我们已经证明了 TIMS 与高性能傅里叶变换离子回旋共振 (FTICR) MS 成功耦合 用于分离和鉴定聚糖键异构体的仪器。在此，我们建议修改TIMS 设备及其控制软件，以提高迁移率分辨率、增加 m/z 操作范围以及更好的 与 ExD-FTICR MS/MS 分析集成。然后我们将利用改进的 TIMS-ExD 方法进行详细的 聚糖的结构表征。我们还将使用 TIMS-ExD MS/MS 与离线 LC 结合使用 分级分离以产生包含已识别的聚糖结构及其碰撞横截面的文库 价值观。该图书馆将向公众开放。 初步开发将在 FTICR MS 平台上进行，因为它提供卓越的质量精度和 分辨率，以及最佳的 ExD 性能。我们在这里开发的技术以后可以 随着替代 ECD 细胞的开发，转移到其他更实惠的 MS 仪器 将 ExD 功能引入非 ICR 仪器。