Immunoglobulin A Carries SulfatedN-glycans Primarily at the Tailpiece Site – An Oxonium-Ion-Guided Approach for Site-SpecificN-glycan Identification

Sulfated N- glycans from human immunoglobulin A (IgA) were recently discovered via glycomic approaches. However, their site-specific description is still pending. Certain N- glycan structures at specific N- glycosylation sites in IgA are crucial for microbial neutralization and effector functions. For instance, sialylated N- glycans on the C-terminal tailpiece mediate anti-viral activity by interfering with sialic-acid-binding viruses. Sulfated N- glycan epitopes can be ligands for viral proteins and thus play a role in the immune response. In this study, we performed a site-specific screening for sulfated N- glycans in two commercially available human serum IgA samples employing an in-depth N- glycoproteomic approach, previously developed by us. We found evidence of complex-type and hybrid-type N- glycans containing sulfated N- acetylhexosamine (sulfated HexNAc) attached to the N- glycosylation sites in the tailpiece and the C H 2 domain of both IgA subclasses. A detailed comparison of the N- glycosylation profiles of human serum IgA samples from two suppliers showed such N- glycans with sulfated HexNAc consistently in higher abundance in the tailpiece region. Surprisingly, also complex-type N- glycan compositions bearing O- acetylated sialic acid were identified in the tailpiece. These findings have not been described before for a site-specific glycopeptide analysis. Overall, our work provides a methodology for performing a dedicated site-specific search for sulfated and O- acetylated N- glycans that can be easily transferred, e.g. to human IgA derived from mucosal tissues, milk, or saliva. Our future aim is to include sulfated N- glycans into longitudinal studies of IgA N- glycosylation and to investigate their role as a biomarker and a treatment option. Abstract Figure