engleski

Fc-linked N-glycosylation of IgG subclasses in three mouse strains analyzed with nanoUPLC-ESI-MS

Biological activity of immunoglobulin G (IgG) is heavily influenced by the biantennary N-glycans attached to the heavy chains of the fragment crystallizable (Fc). The four IgG subclasses recognized in humans and mice differ both in amino acid composition and in their Fc-region N- glycosylation profiles. The aim of the project is to compare N-glycosylation profiles of the Fc- region of IgG subclasses in three commonly used mouse strains. IgG was isolated from the serum of individual sex- and aged-matched mice of BALB/c, C57BL/6 and C3H strains kept in the same environment. The obtained IgG tryptic glycopeptides corresponding to the Fc region of the four mouse IgG subclasses were separated with nano ultra-performance liquid chromatography and quantified with electrospray ionization mass spectrometry (nanoUPLC-ESI-MS) method. We compared several derived N-glycan traits, describing the relative abundance of N- glycan structural features. We observed differences in Fc-linked N-glycosylation profiles both between mouse strains and between IgG subclasses. Interstingly, the C57BL/6 mice carying a rare IgG1 allotype characterized by a Phe -> Ile substitution in the tryptic Fc-glycopeptide showed increased agalactosylation and reduced sialylation of IgG1 compared to BALB/c and C3H strains. Our next goal is to develop a technique for relative quantification of the IgG allotypes in our experimental setup to determine if they are expressed on equal levels in the heterozygous individuals and if their N-glycosylation profiles are different. We plan to further investigate the connection between the abundance of known IgG allotypes and their N-glycosylation profiles, with a particular focus on known IgG1, IgG2b and IgG2c allelic polymorphism in mice and IgG3 in humans, which will require integration of proteomics, genomics and glycomics data.

IgG glycosylation ; mouse ; nanoUPLC-ESI-MS

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