The use of mass spectrometry for the proteomic analysis of glycosylation

Morelle, Willy; Canis, Kevin; Chirat, Frédéric; Faid, Valegh; Michalski, Jean‐Claude

doi:10.1002/pmic.200600129

Cited by 202 publications

(187 citation statements)

References 271 publications

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“…Mass spectrometry and permethylation Electrospray ionization (ESI) and MALDI MS play a crucial role in the characterization of glycosylation 21,22 . MALDI-TOF-MS offers a simple means for screening complex mixtures, and MS-MS methods using electrospray with low-energy collisioninduced dissociation (CID) are valuable tools for the production of fragment ions to investigate the sequence and even linkage of glycans.…”

Section: Release Of Glycansmentioning

confidence: 99%

Analysis of protein glycosylation by mass spectrometry

2007

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We present a detailed protocol for the structural analysis of protein-linked glycans. In this approach, appropriate for glycomics studies, N-linked glycans are released using peptide N-glycosidase F and O-linked glycans are released by reductive alkaline b-elimination. Using strategies based on mass spectrometry (matrix-assisted laser desorption/ionization-time of flight mass spectrometry and nano-electrospray ionization mass spectrometry/mass spectrometry (nano-ESI-MS-MS)), chemical derivatization, sequential exoglycosidase digestions and linkage analysis, the structures of the N-and/or O-glycans are defined. This approach can be used to study the glycosylation of isolated complex glycoproteins or of numerous glycoproteins encountered in a complex biological medium (cells, tissues and physiological fluids).

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Section: Release Of Glycansmentioning

confidence: 99%

Analysis of protein glycosylation by mass spectrometry

2007

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“…Nlinked glycopeptide analysis using typical low-energy CAD is difficult as it generally yields only glycan, but not peptide, structural information. The observation of oxonium ions such as m/z 204 (HexNAc), 163 (Hexose), 292 (sialic acid), and 366 (Hexose-HexNAc) can identify these peptides as glycopeptides (for a more detailed review on the analysis of glycoproteins by mass spectrometry see [35]). …”

Section: O-and N-linked Glycosylationmentioning

confidence: 99%

The utility of ETD mass spectrometry in proteomic analysis

Mikesh

Ueberheide

Chen

et al. 2006

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics

492

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Mass spectrometry has played an integral role in the identification of proteins and their posttranslational modifications (PTM). However, analysis of some PTMs, such as phosphorylation, sulfonation, and glycosylation, is difficult with collision-activated dissociation (CAD) since the modification is labile and preferentially lost over peptide backbone fragmentation, resulting in little to no peptide sequence information. The presence of multiple basic residues also makes peptides exceptionally difficult to sequence by conventional CAD mass spectrometry. Here we review the utility of electron transfer dissociation (ETD) mass spectrometry for sequence analysis of posttranslationally modified and/or highly basic peptides. Phosphorylated, sulfonated, glycosylated, nitrosylated, disulfide bonded, methylated, acetylated, and highly basic peptides have been analyzed by CAD and ETD mass spectrometry. CAD fragmentation typically produced spectra showing limited peptide backbone fragmentation. However, when these peptides were fragmented using ETD, peptide backbone fragmentation produced a complete or almost complete series of ions and thus extensive peptide sequence information. In addition, labile PTMs remained intact. These examples illustrate the utility of ETD as an advantageous tool in proteomic research by readily identifying peptides resistant to analysis by CAD. A further benefit is the ability to analyze larger, non-tryptic peptides, allowing for the detection of multiple PTMs within the context of one another.

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“…Analysis of protein glycosylation by mass spectrometry is typically achieved by two main approaches: The glycans can be released from the peptide backbone either enzymatically or chemically, or the glycoprotein can be subjected to a protease digestion, producing a mixture of peptides and glycopeptides. The latter approach is advantageous to releasing glycans from the protein, since it does not require extra sample manipulation and allows for site-specific glycosylation profiling [18]. However, there are several obstacles encountered when using a glycopeptidebased MS analysis.…”

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confidence: 99%

Comparison of HPLC/ESI-FTICR MS versus MALDI-TOF/TOF MS for glycopeptide analysis of a highly glycosylated HIV envelope glycoprotein

Irungu

Zhang

et al. 2008

J. Am. Soc. Mass Spectrom.

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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⌬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. (J

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The use of mass spectrometry for the proteomic analysis of glycosylation

Cited by 202 publications

References 271 publications

Analysis of protein glycosylation by mass spectrometry

Analysis of protein glycosylation by mass spectrometry

The utility of ETD mass spectrometry in proteomic analysis

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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