A peptide from the staphylococcal protein Efb binds P-selectin and inhibits the interaction of platelets with leukocytes.

Aims: P-selectin is a key surface adhesion molecule for the interaction of platelets with leukocytes. We have shown previously that the N-terminal domain of Staphylococcus aureus extracellular fibrinogen-binding protein (Efb) binds to P-selectin and interferes with platelet-leukocyte aggregate formation. Here, we aimed to identify the minimal Efb motif required for binding platelets and to characterize its ability to interfering with the formation of platelet-leukocyte aggregates.

Methods And Results: Using a library of synthetic peptides, we mapped the platelet-binding site to a continuous 20 amino acid stretch. The peptide Efb was able to bind to resting and, to a greater extent, thrombin-stimulated platelets in the absence of fibrinogen. Dot blots, pull-down assays and P-selectin glycoprotein ligand-1 (PSGL-1) competitive binding experiments identified P-selectin as the cellular docking site mediating Efb platelet binding. Accordingly, Efb did not bind to other blood cells and captured platelets from human whole blood under low shear stress conditions. Efb did not affect platelet activation as tested by aggregometry, flow cytometry and immunoblotting, but inhibited the formation of platelet-leukocyte aggregates (PLAs). Efb also interfered with the platelet-dependent stimulation of neutrophil extracellular traps (NETs) formation in vitro.

Conclusions: We have identified Efb as a novel selective platelet-binding peptide. Efb binds directly to P-selectin and inhibits interactions of platelets with leukocytes that lead to PLA and NET formation. As PLAs and NETs play a key role in thromboinflammation, Efb is an exciting candidate for the development of novel selective inhibitors of the proinflammatory activity of platelets.