質譜分析技術之應用於醣質體學

Mass Spectrometry-based Glycomics and Glycosylation Analysis

林佳葳(Chia-Wei Lin) ； 吳思緯(Sz-Wei Wu) ； 于心宜(Shin-Yi Yu) ； 邱繼輝(Kay-Hooi Khoo)

《化學》 65卷2期 (2007/06) Pp. 125-136

https://doi.org/10.6623/chem.2007014

摘要

高靈敏度質譜分析，搭配各式分離層析、化學衍生與醣水解酶的降解，是鑑定醣類結構的關鍵技術。近年來，單一醣蛋白上的醣質結構分析已延伸到醣質體學上的應用，其研究包含了如何描繪出細胞、組織以至整個生物體的醣質體特徵與其在生理上或是遺傳上的改變。本實驗室多年來致力研發以基質輔助雷射脫附游離法為主，針對甲基化衍生醣質快速進行一次質譜掃描與二次質譜解序的方式，作為初步鑑定醣質體圖譜的實驗策略。近年來，分別藉由Q-TOF與TOF/TOF串聯式質譜儀提供的高低能量碰撞誘導解離法的優缺點與互補性，建立一個可以有效分析複雜醣質結構的斷裂碎片模式，經由低能量碰撞誘導解離法較簡單的圖譜可得整個醣質序列、分支模式、核心結構以及末端結構，進而系統化的從較複雜的高能量碰撞誘導解離斷裂碎片，得到鏈結位置的資訊。本文主要是簡述此一醣質體分析法，強調需要對醣生化學有一個最基本的認識及樣品前處理和化學衍生的重要性，方能有效的判讀二次質譜數據，成功的取得豐富的醣質體資訊，進一步推動功能性醣蛋白質體學的研究。

關鍵字

醣化 ； 醣質體 ； 醣序列分析 ； 質譜儀 ； 時間飛行式串聯質譜儀

並列摘要

Mass spectrometry (MS) and various modes of separation techniques, coupled with chemical modifications and glycosidase digestions, are the corner stones of high sensitivity approach in structural determination of complex glycans derived from biological source. More recently, these glycosylation studies have evolved into what is referred to as glycomics which involves mapping the complexity and characteristics of the glycome of a cell, tissue, or organism, at a particular physiological or genetically altered state. From the onset, we and others have advocated high throughput and robust MALDI-based MS profiling and CID MS/MS sequencing of permethyl derivatives as the principal ＂first screen＂ strategy in glycomics. Over the last few years, an extensive library of both low and high energy CID MALDI MS/MS fragmentation pattern encompassing a full range of complex glycans, as acquired on the Q/TOF and TOF/TOF, respectively, have been established. These studies expose the relative strengths and limitations of the highly complementary low and high energy CID MS/MS. While the overall sequence, branching pattern, core type and terminal epitope substituents were found to be best determined via low energy CID MS/MS due to its simplicity, high energy CID MS/MS on a TOF/TOF can provide linkage information once a systematic assignment of the complicated cleavage pattern has been established. Either this or the multistage MSn approach on ion traps can thus provide an information rich glycomic map, from which further targeted glycoproteomics and functional studies can be designed. This article aims to give a brief account of our own experience in MALDI-MS-based glycomics while emphasizing the need for a basic understanding of current glyco-structure repertoire and glycobiology, so as to facilitate MS/MS data interpretation. Equally important is the requisite sample preparation and chemical derivatization for a better experience and success in MS analyses of complex glycans.

並列關鍵字

Glycosylation ； Glycomics ； Glycan sequencing ； Mass spectrometry ； TOF/TOF