Structural Analysis of GPI-glycans from GPI-anchored Proteins by Mass Spectrometry

Glycosylphosphatidylinositol-anchored proteins (GPI-APs) are a group of proteins anchored to glycolipid on the cell membrane and are ubiquitous in eukaryotes. The basic structure of the glycosylphosphatidylinositol (GPI) moiety comprises ethanolamine phosphate, three mannose residues, glucosamine and phosphatidylinositol (PI). This basic structure and the mechanism by which proteins are anchored to membranes via the structure are conserved among organisms. After the identification of paroxysmal nocturnal hemoglobinuria (PNH), diseases derived from GPI anchor deficiencies were discovered. To comprehend these diseases fully, a comprehensive understanding of GPI anchor biosynthesis, encompassing the intricate remodeling of glycans and lipids, becomes imperative. We used mutant strains of Saccharomyces cerevisiae in which the gene encoding the enzyme that catalyzes remodeling was knocked out and a model molecule for GPI-APs to observe the remodeling process. Herein, we introduce a method for analyzing and identifying glycan structures in GPI anchors using liquid chromatography-electrospray ionization mass spectrometry (LC-ESI MS).