Comparative Analysis of Different N-glycan Preparation Approaches and Development of Optimized Solid-Phase Permethylation Using Mass Spectrometry

Guan，Yudong1; Zhang，Min2; Wang，Jigang1; Schluter，Hartmut2

2021

10.1021/acs.jproteome.1c00135

JOURNAL OF PROTEOME RESEARCH   影响因子和分区

1535-3893

1535-3907

Protein N-glycosylation characterization is challenging due to structural micro- and macro-heterogeneity. Although various N-glycan preparation strategies, including purification and derivatization, have been previously developed prior to mass spectrometric analysis, systematic evaluation still needs to be performed. This study compared the different N-glycan purification strategies, including filter-aided sample preparation, de-N-glycosylated protein precipitation, and trypsin digestion followed by reversed phase-based solid-phase extraction, and derivatization approaches, such as solid-phase permethylation, reductive amination, and reduction. With the comparative analysis, an optimized solid-phase permethylation (OSPP) workflow was developed for mass spectrometric N-glycomics, showing simplified analysis for N-glycan compositions and high yields using etanercept. The N-glycan samples released from trastuzumab and adalimumab were utilized to test OSPP to obtain their N-glycan profiles using mass spectrometry. Based on different standard procedures across laboratories, this study provides the reference for analysts to select an appropriate N-glycan preparation method with their research purposes.

mass spectrometry N-glycan permethylation reduction reductive amination

SCI

英语

第一 ; 通讯

Program of the China Scholarship Council (CSC)[201606220045,201806780022]

Biochemistry & Molecular Biology

Biochemical Research Methods

WOS:000649269600065

AMER CHEMICAL SOC

BIOLOGY & BIOCHEMISTRY

2-s2.0-85104918064

Scopus

1.The First Affiliated Hospital (Shenzhen People's Hospital),Southern University of Science and Technology,Shenzhen,518055,China
2.Institute of Clinical Chemistry,Laboratory Medicine,University Medical Center Hamburg-Eppendorf,Hamburg,20246,Germany

Guan，Yudong,Zhang，Min,Wang，Jigang,et al. Comparative Analysis of Different N-glycan Preparation Approaches and Development of Optimized Solid-Phase Permethylation Using Mass Spectrometry[J]. JOURNAL OF PROTEOME RESEARCH,2021,20(5).

Guan，Yudong,Zhang，Min,Wang，Jigang,&Schluter，Hartmut.(2021).Comparative Analysis of Different N-glycan Preparation Approaches and Development of Optimized Solid-Phase Permethylation Using Mass Spectrometry.JOURNAL OF PROTEOME RESEARCH,20(5).

Guan，Yudong,et al."Comparative Analysis of Different N-glycan Preparation Approaches and Development of Optimized Solid-Phase Permethylation Using Mass Spectrometry".JOURNAL OF PROTEOME RESEARCH 20.5(2021).