Enhanced Quantitative LC-MS/MS Analysis of N‑linked Glycans Derived from Glycoproteins Using Sodium Deoxycholate Detergent

Protein glycosylation is a common protein post-translational modification (PTM) in living organisms and has been shown to associate with multiple diseases, and thus may potentially be a biomarker of such diseases. Efficient protein/glycoprotein extraction is a crucial step in the preparation of N-gl...

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Published inJournal of proteome research Vol. 17; no. 8; pp. 2668 - 2678
Main Authors Zhu, Rui, Zhou, Shiyue, Peng, Wenjing, Huang, Yifan, Mirzaei, Parvin, Donohoo, Kaitlyn, Mechref, Yehia
Format Journal Article
LanguageEnglish
Published United States American Chemical Society 03.08.2018
Subjects
Animals
Cell Line, Tumor
Chemical Precipitation
Chromatography, Liquid - methods
Deoxycholic Acid - chemistry
Glycoproteins - analysis
Glycosylation
Humans
Polysaccharides - analysis
Protein Processing, Post-Translational
Proteomics - methods
Reproducibility of Results
Specimen Handling
Tandem Mass Spectrometry - methods
sodium deoxycholate
acidic labile detergent
detergent removal
N-glycomics
LC-MS/MS sample preparation
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Summary:Protein glycosylation is a common protein post-translational modification (PTM) in living organisms and has been shown to associate with multiple diseases, and thus may potentially be a biomarker of such diseases. Efficient protein/glycoprotein extraction is a crucial step in the preparation of N-glycans derived from glycoproteins prior to LC-MS analysis. Convenient, efficient and unbiased sample preparation protocols are needed. Herein, we evaluated the use of sodium deoxycholate (SDC) acidic labile detergent to release N-glycans of glycoproteins derived from biological samples such as cancer cell lines. Compared to the filter-aided sample preparation approach, the sodium deoxycholate (SDC) assisted approach was determined to be more efficient and unbiased. SDC removal was determined to be more efficient when using acidic precipitation rather than ethyl acetate phase transfer. Efficient extraction of proteins/glycoproteins from biological samples was achieved by combining SDC lysis buffer and beads beating cell disruption. This was suggested by a significant overall increase in the intensities of N-glycans released from cancer cell lines. Additionally, the use of SDC approach was also shown to be more reproducible than those methods that do not use SDC.
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ISSN:1535-3893
1535-3907
DOI:10.1021/acs.jproteome.8b00127