Comparison of HPLC/ESI-FTICR MS versus MALDI-TOF/TOF MS for glycopeptide analysis of a highly glycosylated HIV envelope glycoprotein
Comparison of HPLC/ESI-FTICR MS versus MALDI-TOF/TOF MS for glycopeptide analysis of a highly glycosylated HIV envelope glycoprotein
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DOI：
10.1016/j.jasms.2008.05.010
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发表时间：
2008-08-01
影响因子：
3.2
通讯作者：
Desaire, Heather
中科院分区：
文献类型：
--
作者：
Irungu, Janet;Go, Eden P.;Desaire, Heather
确定分泌蛋白和病毒包膜蛋白上聚糖的结构和位置对于理解糖基化如何影响其生物学特性非常重要。基于糖肽质谱（MS）的分析是一种非常强大的新兴方法来表征糖蛋白，其中糖基化位点和相应的聚糖结构在单个MS实验中得到阐明。然而，迄今为止，关于哪种质谱平台提供最佳糖基化覆盖信息还没有达成共识。在此，我们采用了两种最广泛使用的质谱方法，在线高效液相色谱-电喷雾电离质谱（HPLC/ESI-MS）和离线HPLC，随后基质辅助激光解吸/电离质谱（MALDI-MS），以确定两种方法中的哪一种提供了复合糖蛋白（M组共有HIV-1包膜）的最佳糖基化覆盖信息，CON-S gp 140 Delta CFI，其具有31个潜在的糖基化位点。我们的研究结果突出了两种方法之间获得的信息内容的差异，如检测到的糖基化位点的总数，每个位点存在的N-连接聚糖的数量，以及使用MS/MS实验获得的关于糖肽的确证性信息的类型。这两种方法在糖肽的覆盖范围和MS/MS实验中提供的信息方面都是非常互补的。在这项研究中的信息有助于质谱领域的优势和局限性，证明了糖蛋白分析中两个广泛使用的MS平台。
Defining the structures and locations of the glycans attached on secreted proteins and virus envelope proteins is important in understanding how glycosylation affects their biological properties. Glycopeptide mass spectrometry (MS)-based analysis is a very powerful, emerging approach to characterize glycoproteins, in which glycosylation sites and the corresponding glycan structures are elucidated in a single MS experiment. However, to date there is not a consensus regarding which mass spectrometric platform provides the best glycosylation coverage information. Herein, we employ two of the most widely used MS approaches, online high performance liquid chromatography-electrospray ionization mass spectrometry (HPLC/ESI-MS) and offline HPLC followed by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), to determine which of the two approaches provides the best glycosylation coverage information of a complex glycoprotein, the group M consensus HIV-1 envelope, CON-S gp140 Delta CFI, which has 31 potential glycosylation sites. Our results highlight differences in the informational content obtained between the two methods such as the overall number of glycosylation sites detected, the numbers of N-linked glycans present at each site, and the type of confirmatory information obtained about the glycopeptide using MS/MS experiments. The two approaches are quite complementary, both in their coverage of glycopeptides and in the information they provide in MS/MS experiments. The information in this study contributes to the field of mass spectrometry by demonstrating the strengths and limitations of two widely used MS platforms in glycoprotein analysis.