Structures of the O-glycans on P-selectin glycoprotein ligand-1 from HL-60 cells

P-selectin glycoprotein ligand-1 (PSGL-1) is a disulfide-bonded homodimeric mucin-like glycoprotein on leukocytes that interacts with both P- and E-selectin. In this report we describe the structures of the Ser/Thr-linked O-glycans of PSGL-1 synthesized by HL-60 cells metabolically radiolabeled with H-3-sugar precursors. In control studies, the O-glycans on CD43 (leukosialin), a mucin-like glycoprotein also expressed by HL 60 cells, were analyzed and compared to those of PSGL-1, O-Glycans were released from Ser/Thr residues by mild base/borohydride treatment of purified glycoproteins, and glycan structures were determined by a combination of techniques, In contrast to expectations, PSGL-1 is not heavily fucosylated; a majority of the O-glycans are disialylated or neutral forms of the core-2 tetrasaccharide GaI beta 1-4GlcNAc beta 1-6(Gal beta 1-->3)GaINAcOH, A minority of the O-glycans are alpha-1,3-fucosylated that occur as two major species containing the sialyl Lewis x antigen; one species is a disialylated, monofucosylated glycan, [GRAPHICS] and the other is a monosialylated, trifucosylated glycan having a polylactosamine backbone. [GRAPHICS] CD43 lacks the fucosylated glycans found on PSGL-1 and is enriched for the nonfucosylated, disialylated core-2 hexasaccharide. These results demonstrate that PSGL-1 contains unique fucosylated O-glycans that are predicted to be critical for high affinity interactions between PSGL-1 and selectins.