Binding of human serum amyloid P componentto L‐selectin

Abstract

Serum concentrations of soluble L‐selectin by far exceed those of other soluble adhesion molecules, and serum soluble L‐selectin concentrations are remarkably stable upon prolonged storage. We present evidence for Ca²⁺‐dependent binding interactions between human serum amyloid P (SAP), a proteolysis‐resistant pentraxin glycoprotein, and L‐selectin, as shown by surface plasmon resonance measurements, protein band shift assays in a native PAGE system, and after SDS‐PAGE and membrane transfer. Monoclonal antibodies to L‐selectin strongly reduced binding of biotinylated SAP to L‐selectin‐IgG chimeras immobilized on microtiter plates. As binding was reduced by prior glycopeptidase F treatment of L‐selectin but not of SAP, it appears to be based on SAP lectin domain interactions with N‐linked L‐selectin carbohydrates. In freshly prepared human lymphocytes, SAP incubation induced expression of a β2 integrin neoepitope associated with high‐affinity binding. This was partially blocked by pre‐incubation with Fab fragments of two anti‐L‐selectin antibodies. In flow chamber experiments, SAP inhibited the adherence of human neutrophils to activated endothelium under shear stress. Thus, SAP binds to human L‐selectin and affects L‐selectin‐dependent leukocyte‐endothelial interactions.