A FUNCTIONAL EPITOPE ON P-SELECTIN THAT SUPPORTS BINDING OF P-SELECTIN TO P-SELECTIN GLYCOPROTEIN LIGAND-1 BUT NOT TO SIALYL-LEWIS-X OLIGOSACCHARIDES

P-selectin mediates the adhesion of leukocytes to activated platelets and endothelial cells. To characterize the functional domains of P-sel ectin for ligand recognition, we established nine hybridoma cell lines secreting anti-rat P-selectin mAb, Among them, the mAb C215 bound bot h rat and human P-selectins, and inhibited binding of rat and human P- selectins to P-selectin glycoprotein ligand-1 (PSGL-1) from HL-60 cell s. In contrast, mAb C215 failed to inhibit the binding of rat and huma n P-selectin-IgG to sialyl Lewis X (sLe(X)) oligosaccharides. Epitope mapping of mAb C215 using synthetic decapeptides revealed that mAb C21 5 binds specifically to an eight-residue epitope that spans amino acid s 76-83 of rat P-selectin, a region completely conserved by human P-se lectin. Synthetic peptides containing the mAb C215 epitope inhibited b inding of P-selectin to PSGL-1, but not to sLe(X) oligosaccharides, su ggesting that the C215 epitope on P-selectin may directly interact wit h a particular site on the PSGL-1. core protein essential for interact ion with P-selectin, such as sulfated tyrosine residues. Our results s uggest the presence of two ligand recognition sites on P-selectin nece ssary for binding to PSGL-1-one recognizes sLe(X), while the other rec ognises the PSGL-1 core protein.