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

Int Immunol. 1998 May;10(5):639-49. doi: 10.1093/intimm/10.5.639.

Abstract

P-selectin mediates the adhesion of leukocytes to activated platelets and endothelial cells. To characterize the functional domains of P-selectin for ligand recognition, we established nine hybridoma cell lines secreting anti-rat P-selectin mAb. Among them, the mAb C215 bound both 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 cells. In contrast, mAb C215 failed to inhibit the binding of rat and human P-selectin-IgG to sialyl Lewis X (sLe(x)) oligosaccharides. Epitope mapping of mAb C215 using synthetic decapeptides revealed that mAb C215 binds specifically to an eight-residue epitope that spans amino acids 76-83 of rat P-selectin, a region completely conserved by human P-selectin. Synthetic peptides containing the mAb C215 epitope inhibited binding of P-selectin to PSGL-1, but not to sLe(x) oligosaccharides, suggesting that the C215 epitope on P-selectin may directly interact with a particular site on the PSGL-1 core protein essential for interaction with P-selectin, such as sulfated tyrosine residues. Our results suggest the presence of two ligand recognition sites on P-selectin necessary for binding to PSGL-1--one recognizes sLe(x), while the other recognises the PSGL-1 core protein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Antibodies, Monoclonal
  • Binding Sites
  • Cell Adhesion
  • Epitope Mapping
  • Epitopes / genetics
  • Epitopes / metabolism
  • HL-60 Cells
  • Humans
  • Immunoglobulin G / metabolism
  • In Vitro Techniques
  • Lewis Blood Group Antigens
  • Ligands
  • Membrane Glycoproteins / metabolism*
  • Mice
  • Molecular Sequence Data
  • Oligosaccharides / metabolism*
  • P-Selectin / genetics
  • P-Selectin / immunology*
  • P-Selectin / metabolism*
  • Protein Binding
  • Rats
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / immunology
  • Recombinant Fusion Proteins / metabolism
  • Sialyl Lewis X Antigen

Substances

  • Antibodies, Monoclonal
  • Epitopes
  • Immunoglobulin G
  • Lewis Blood Group Antigens
  • Ligands
  • Membrane Glycoproteins
  • Oligosaccharides
  • P-Selectin
  • P-selectin ligand protein
  • Recombinant Fusion Proteins
  • Sialyl Lewis X Antigen