Document Detail


IgG Fab glycosylation analysis using a new mass spectrometric high-throughput profiling method reveals pregnancy-associated changes.
MedLine Citation:
PMID:  25004930     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
The N-linked glycosylation of the constant fragment (Fc) of Immunoglobulin G has been shown to change during pathological and physiological events and to strongly influence antibody inflammatory properties. In contrast, little is known about Fab-linked N-glycosylation carried by approximately 20% of IgG. Here we present a high-throughput workflow to analyze Fab and Fc glycosylation of polyclonal IgG purified from 5 μL serum. Thirty-seven different N-glycans could be detected and quantified by MALDI-TOF-MS analysis in reflectron positive mode using a novel linkage specific derivatization of sialic acid. This method was applied to 174 samples of a pregnancy cohort to reveal Fab glycosylation features and their change with pregnancy. Data analysis revealed marked differences between Fab and Fc glycosylation, especially in the levels of galactosylation, sialylation, incidence of bisecting GlcNAc and presence of high mannose structures, which are all higher in the Fab portion compared to Fc, while Fc showed higher levels of fucosylation. Additionally, we observed several changes during pregnancy and after delivery. Fab N-glycan sialylation was increased and bisection decreased as compared to post-partum time points, while nearly complete galactosylation of Fab glycans was observed throughout. Fc glycosylation changes were similar to results described before, with increased galactosylation and sialylation and decreased bisection during pregnancy. We expect that the parallel analysis of IgG Fab and Fc, as set up in this paper, will be important for unraveling roles of these glycans in (auto)immunity, which may be mediated via recognition by human lectins or modulation of antigen binding.
Authors:
Albert Bondt; Yoann Rombouts; Maurice H J Selman; Paul J Hensbergen; Karli R Reiding; Johanna M W Hazes; Radboud J E M Dolhain; Manfred Wuhrer
Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2014-7-8
Journal Detail:
Title:  Molecular & cellular proteomics : MCP     Volume:  -     ISSN:  1535-9484     ISO Abbreviation:  Mol. Cell Proteomics     Publication Date:  2014 Jul 
Date Detail:
Created Date:  2014-7-9     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  101125647     Medline TA:  Mol Cell Proteomics     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Copyright Information:
Copyright © 2014, The American Society for Biochemistry and Molecular Biology.
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