Site-specific glycosylation mapping of Fc gamma receptor IIIb from neutrophils of health donors

Abstract

Fc gamma receptors (Fc{gamma}R) translate antigen-recognition by immunoglobulin G (IgG) into various immune responses. A better understanding of this key element of immunity promises novel insights into mechanisms of (auto-/allo-)immune diseases and more rationally designed antibody-based drugs. Glycosylation on both IgG and Fc{gamma}R impacts their interaction dramatically. In this study, we developed a straightforward and comprehensive analytical methodology to map Fc{gamma}RIIIb glycosylation from primary human material. In contrast to recently published alternatives, we assessed all glycosylation sites in a single LC-MS/MS run and simultaneously determined the donor allotype. Studying Fc{gamma}RIIIb derived from healthy donor neutrophils, we observed profound differences as compared to the soluble variant and the homologous Fc{gamma}RIIIa on natural killer cells. This method will allow assessment of Fc{gamma}RIII glycosylation differences between individuals, cell types, subcellular locations and pathophysiological conditions.