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Evaluation and Optimization of High-Field Asymmetric Waveform Ion-Mobility Spectrometry for Multiplexed Quantitative Site-Specific N-Glycoproteomics
- Source :
- Analytical Chemistry. 93:8846-8855
- Publication Year :
- 2021
- Publisher :
- American Chemical Society (ACS), 2021.
-
Abstract
- The heterogeneity and complexity of glycosylation hinder the depth of site-specific glycoproteomics analysis. High-field asymmetric-waveform ion-mobility spectrometry (FAIMS) has shown to improve the scope of bottom-up proteomics. The benefits of FAIMS for quantitative N-glycoproteomics have not been investigated yet. In this work, we optimized FAIMS settings for N-glycopeptide identification, with or without the tandem mass tag (TMT) label. The optimized FAIMS approach significantly increased the identification of site-specific N-glycopeptides derived from the purified IgM protein or human lymphoma cells. We explored in detail the changes in FAIMS mobility caused by N-glycopeptides with different characteristics, including TMT labeling, charge state, glycan type, peptide sequence, glycan size and precursor m/z. Importantly, FAIMS also improved multiplexed N-glycopeptide quantification, both with the standard MS2 acquisition method and with our recently developed Glyco-SPS-MS3 method. The combination of FAIMS and Glyco-SPS-MS3 provided the highest quantitative accuracy and precision. Our results demonstrate the advantages of FAIMS for improved mass-spectrometry-based qualitative and quantitative N-glycoproteomics. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=75 SRC="FIGDIR/small/436434v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@126b92forg.highwire.dtl.DTLVardef@147d6e5org.highwire.dtl.DTLVardef@16d6eb0org.highwire.dtl.DTLVardef@17dfe2a_HPS_FORMAT_FIGEXP M_FIG C_FIG
- Subjects :
- Glycan
Chromatography
biology
010405 organic chemistry
Ion-mobility spectrometry
Chemistry
010401 analytical chemistry
010402 general chemistry
Mass spectrometry
Tandem mass tag
Proteomics
01 natural sciences
Multiplexing
0104 chemical sciences
Analytical Chemistry
Glycoproteomics
biology.protein
High field
Asymmetric waveform
Subjects
Details
- ISSN :
- 15206882 and 00032700
- Volume :
- 93
- Database :
- OpenAIRE
- Journal :
- Analytical Chemistry
- Accession number :
- edsair.doi.dedup.....c1f79a838db864e614f08a67ff310a1e
- Full Text :
- https://doi.org/10.1021/acs.analchem.1c00802