Title : Site-specific N-glycosylation analysis of human
immunoglobulin e .
Immunoglobulin E (
IgE ) is a heterodimeric
glycoprotein involved in
antiparasitic and allergic immune reactions
Abstract :
- IgE glycosylation is known to exhibit significant interindividual variation, and several reports have indicated its relevance in determining IgE activity
- Here, we present site-specific glycosylation analysis of IgE from three different sources: IgE from the serum of a hyperimmune donor, from the pooled serum of multiple nondiseased donors, and from the pooled serum of 2 patients with IgE myeloma
- The heavy chains were isolated and digested with either trypsin, proteinase K , or chymotrypsin, which permitted coverage of all seven potential N-glycosylation sites.
- The resulting (glyco-) peptides were analyzed by nano-reversed-phase-LC-MS/MS and MALDI-TOF/TOF-MS /MS
- Site Asn264 was shown to be unoccupied
- In all three samples, site Asn275 contained exclusively oligomannosidic structures with between 2 and 9 mannoses, whereas sites Asn21, Asn49, Asn99, Asn146, and Asn252 contained exclusively complex-type glycans
- For the nonmyeloma IgE , the majority of these glycans were biantennary and core-fucosylated and contained one or two terminal N-acetylneuraminic acids
- In contrast, myeloma IgE showed a higher abundance of triantennary and tetraantennary glycan structures and a low abundance of species with a bisecting N-acetylglucosamine.
- Our approach allows comparison of the glycosylation of IgE samples in a site-specific manner