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

Stuart Wallis, Nina Wolska, Hanna Englert, Mareike Posner, Abhishek Upadhyay, Thomas Renne, Ian Eggleston, Stefan Bagby, Giordano Pula

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Abstract

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 S. 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 characterise 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 Efb68-87 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 Efb68-87 platelet binding. Accordingly, Efb68-87 did not bind to other blood cells and captured platelets from human whole blood under low shear stress conditions. Efb68-87 did not affect platelet activation as tested by aggregometry, flow cytometry and immunoblotting, but inhibited the formation of platelet-leukocyte aggregates (PLAs). Efb68-87 also interfered with the platelet-dependent stimulation of neutrophil extracellular traps (NETs) formation in vitro.

Conclusions
We have identified Efb68-87 as a novel selective platelet-binding peptide. Efb68-87 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, Efb68-87 is an exciting candidate for the development of novel selective inhibitors of the proinflammatory activity of platelets.
Original languageEnglish
Pages (from-to)729-741
Number of pages13
JournalJournal of Thrombosis and Haemostasis
Volume20
Issue number3
Early online date30 Nov 2021
DOIs
Publication statusPublished - 31 Mar 2022

Bibliographical note

Funding Information:
The authors thank the DFG‐funded Cytometry and Cell Sorting Core Unit at the UKE hospital (Hamburg). This work was funded by the British Heart Foundation for the financial support of SW, SB and GP (FS/17/13/3269), the Werner Otto Foundation for the financial support of NW and GP (BN3/97) and the European Research Council for the support of GP (EU project 101025074). Open access funding enabled and organized by ProjektDEAL.

Keywords

  • leukocyte
  • neutrophil extracellular trap
  • P-selectin
  • platelet
  • platelet-leukocyte aggregate
  • thromboinflammation

ASJC Scopus subject areas

  • Hematology

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