An activated L-selectin mutant with conserved equilibrium binding properties but enhanced ligand recognition under shear flow.
Selectins mediate the initial tethering and rolling of leukocytes on vessel walls. Adhesion by selectins is a function of both ligand recognition at equilibrium and mechanical properties of the selectin-ligand bond under applied force. We describe an EGF domain mutant of L-selectin with profoundly augmented adhesiveness over that of native L-selectin but conserved ligand specificity. This mutant, termed LPL, was derived by a substitution of the EGF-like domain of L-selectin with the homologous domain from P-selectin. The mutant bound soluble carbohydrate L-selectin ligand with affinity comparable with that of native L-selectin but interacted with all surface-bound ligands much more readily than native L-selectin, in particular under elevated shear flow. Tethers mediated by both native and mutant L-selectin exhibited similar lifetimes under a range of shear stresses, but the rate of bond formation by the mutant was at least 10-fold higher than that of native L-selectin toward distinct L-selectin ligands. Enhanced rate of bond formation by the mutant was associated with profoundly stronger rolling interactions and reduced dependence of rolling on a threshold of shear stress. This is the first demonstration that the EGF domain can modulate the binding of the lectin domain of a selectin to surface-immobilized ligands under shear flow without affecting the equilibrium properties of the selectin toward soluble ligands.
Dwir
O
O
Department of Immunology, Weizmann Institute of Science, Rehovot, 76100 Israel.
Kansas
G S
GS
Alon
R
R
D000818:Animals; D002460:Cell Line; D004815:Epidermal Growth Factor / Q000235:genetics / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D007700:Kinetics; D019041:L-Selectin / Q000235:genetics*; D008024:Ligands; D051379:Mice; D009154:Mutation; D011485:Protein Binding
D008024:Ligands; D019041:L-Selectin; D004815:Epidermal Growth Factor