Differential glycosylation of polyclonal IgG, IgG-Fc and IgG-Fab isolated from the sera of patients with ANCA-associated systemic vasculitis

Holland, Mark J. G.; Yagi, Hirokazu; Takahashi, Nobutaka; Kato, Koichi; Savage, Caroline O. S.; Goodall, D.M.; Jefferis, Royston

doi:10.1016/j.bbagen.2005.11.021

Cited by 169 publications

(147 citation statements)

References 38 publications

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“…Indeed, it is well established that all IgG carry a conserved N-linked glycosylation site at Asn297, in the CH2 domain of the constant region [32]. In addition, 15-20% of human serum IgG also bear complex diantennary oligosaccharides attached to the variable regions of their light or heavy chains [33,34]. PHA binds preferentially to galactosylated complex diantennary oligosaccharides [35], and this glycan structure is found mainly on the Fab region of human serum IgG in contrast to oligosaccharides in the Fc region which are hypogalactosylated [34].…”

Section: Discussionmentioning

confidence: 99%

“…The IVIg-PHA fraction contained about 80% of the total amount of IgG loaded onto the columns (14·6 mg recovered from 20 mg of IVIg loaded onto the column), indicating that at least 20% of the IgG present in IVIg were bound to or retarded in the PHA-Sepharose column. This amount of lectin-reactive IgG corresponds to the proportion of Fab-glycosylated IgG in human serum [33,34], suggesting that glycans present in the IgG variable region rather than polyreactive IgG are responsible for binding and neutralizing PHA.…”

Section: Discussionmentioning

confidence: 99%

“…In addition, 15-20% of human serum IgG also bear complex diantennary oligosaccharides attached to the variable regions of their light or heavy chains [33,34]. PHA binds preferentially to galactosylated complex diantennary oligosaccharides [35], and this glycan structure is found mainly on the Fab region of human serum IgG in contrast to oligosaccharides in the Fc region which are hypogalactosylated [34]. This suggests that PHA would bind efficiently to glycans present in the Fab portion but not in the Fc region of the IgG.…”

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Neutralization of mitogenic lectins by intravenous immunoglobulin (IVIg) prevents T cell activation: does IVIg really have a direct effect on T cells?

Padet

St‐Amour

Aubin

et al. 2011

Clinical and Experimental Immunology

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Neutralization of mitogenic lectins by intravenous immunoglobulin (IVIg) prevents T cell activation: does IVIg really have a direct effect on T cells?

Padet

St‐Amour

Aubin

et al. 2011

Clinical and Experimental Immunology

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“…Increased levels of these glycosylation features observed in the HILIC analysis may largely be linked to the inclusion of Fab glycans, as Fab glycans are known to show an increased incidence of fully galactosylated, sialylated, and bisected structures compared to the Fc. 12,14,15,36 However, the analysis of sialylation levels by mass spectrometry is complicated by two phenomena: first, the charge introduced by the sialic acid will influence ionization. It may be assumed that negative-mode ionization of the sialylated species was more efficient than the ionization of glycopeptides with nonsialylated, neutral glycan chains.…”

Section: Journal Of Proteome Researchmentioning

confidence: 99%

High-Throughput IgG Fc N-Glycosylation Profiling by Mass Spectrometry of Glycopeptides

et al. 2013

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Age and sex dependence of subclass specific immunoglobulin G (IgG) Fc N-glycosylation was evaluated for 1709 individuals from two isolated human populations. IgGs were obtained from plasma by affinity purification using 96-well protein G monolithic plates and digested with trypsin. Fc Nglycopeptides were purified and analyzed by negative-mode MALDI-TOF-MS with 4-chloro-α-cyanocinnamic acid (Cl-CCA) matrix. Age-associated glycosylation changes were more pronounced in younger individuals (<57 years) than in older individuals (>57 years) and in females than in males. Galactosylation and sialylation decreased with increasing age and showed significant sex dependence. Interestingly, the most prominent drop in the levels of galactosylated and sialylated glycoforms in females was observed around the age of 45 to 60 years when females usually enter menopause. The incidence of bisecting N-acetylglucosamine increased in younger individuals and reached a plateau at older age. Furthermore, we compared the results to the total IgG N-glycosylation of the same populations recently analyzed by hydrophilic interaction liquid chromatography (HILIC). Significant differences were observed in the levels of galactosylation, bisecting N-acetylglucosamine and particularly sialylation, which were shown to be higher in HILIC analysis. Age and sex association of glycosylation features was, to a large extent, comparable between MALDI-TOF-MS and HILIC IgG glycosylation profiling.

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“…Considering the different oligosaccharide profiles of IgG Fab and Fc fragments revealed in some case-control studies [22,23] , we also analyzed the N-glycan profiles of antibody Fab and Fc fragments. Both the Fab and Fc fragments of the bisec-EGFR mAb showed an increased bisecting GlcNAc content compared to the wild type EGFR mAb, but the Fab and Fc fragments had different N-glycan profiles from each other.…”

Section: Wwwchinapharcom Yi Ch Et Almentioning

confidence: 99%

Function characterization of a glyco-engineered anti-EGFR monoclonal antibody cetuximab in vitro

Ruan

Wang

et al. 2014

Acta Pharmacol Sin

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Aim: To evaluate the biochemical features and activities of a glyco-engineered form of the anti-human epidermal growth factor receptor monoclonal antibody (EGFR mAb) cetuximab in vitro. Methods: The genes encoding the Chinese hamster bisecting glycosylation enzyme (GnTIII) and anti-human EGFR mAb were cloned and coexpressed in CHO DG44 cells. The bisecting-glycosylated recombinant EGFR mAb (bisec-EGFR mAb) produced by these cells was characterized with regard to its glycan profile, antiproliferative activity, Fc receptor binding affinity and cell lysis capability. The content of galactose-α-1,3-galactose (α-Gal) in the bisec-EGFR mAb was measured using HPAEC-PAD. Results: The bisec-EGFR mAb had a higher content of bisecting N-acetylglucosamine residues. Compared to the wild type EGFR mAb, the bisec-EGFR mAb exhibited 3-fold higher cell lysis capability in the antibody-dependent cellular cytotoxicity assay, and 1.36-fold higher antiproliferative activity against the human epidermoid carcinoma line A431. Furthermore, the bisec-EGFR mAb had a higher binding affinity for human FcγRIa and FcγRIIIa-158F than the wild type EGFR mAb. Moreover, α-Gal, which was responsible for cetuximab-induced hypersensitivity reactions, was not detected in the bisec-EGFR mAb. Conclusion: The glyco-engineered EGFR mAb with more bisecting modifications and lower α-Gal content than the approved therapeutic antibody Erbitux shows improved functionality in vitro, and requires in vivo validations.

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Differential glycosylation of polyclonal IgG, IgG-Fc and IgG-Fab isolated from the sera of patients with ANCA-associated systemic vasculitis

Cited by 169 publications

References 38 publications

Neutralization of mitogenic lectins by intravenous immunoglobulin (IVIg) prevents T cell activation: does IVIg really have a direct effect on T cells?

Neutralization of mitogenic lectins by intravenous immunoglobulin (IVIg) prevents T cell activation: does IVIg really have a direct effect on T cells?

High-Throughput IgG Fc N-Glycosylation Profiling by Mass Spectrometry of Glycopeptides

Function characterization of a glyco-engineered anti-EGFR monoclonal antibody cetuximab in vitro

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