Inhibition of αIIbβ3 Ligand Binding by an αIIb Peptide that Clasps the Hybrid Domain to the βI Domain of β3.

Wen Hwa Lee,Vassilios Moussis,Dominique Baruch,Elisabeth Schaffner-Reckinger,Vassilios Tsikaris,Paraskevi Trypou,Kelly Aylward,Christilla Bachelot-Loza,Demokritos C Tsoukatos,Marion Egot,Nelly Kieffer

doi:10.1371/journal.pone.0134952

Abstract

Agonist-stimulated platelet activation triggers conformational changes of integrin αIIbβ3, allowing fibrinogen binding and platelet aggregation. We have previously shown that an octapeptide, p1YMESRADR8, corresponding to amino acids 313–320 of the β-ribbon extending from the β-propeller domain of αIIb, acts as a potent inhibitor of platelet aggregation. Here we have performed in silico modelling analysis of the interaction of this peptide with αIIbβ3 in its bent and closed (not swing-out) conformation and show that the peptide is able to act as a substitute for the β-ribbon by forming a clasp restraining the β3 hybrid and βI domains in a closed conformation. The involvement of species-specific residues of the β3 hybrid domain (E356 and K384) and the β1 domain (E297) as well as an intrapeptide bond (pE315-pR317) were confirmed as important for this interaction by mutagenesis studies of αIIbβ3 expressed in CHO cells and native or substituted peptide inhibitory studies on platelet functions. Furthermore, NMR data corroborate the above results. Our findings provide insight into the important functional role of the αIIb β-ribbon in preventing integrin αIIbβ3 head piece opening, and highlight a potential new therapeutic approach to prevent integrin ligand binding.

Highlights

Integrin αIIbβ3, the platelet fibrinogen and von Willebrand factor receptor, plays a key role in hemostasis and thrombosis, by promoting platelet aggregation and thrombus formation at sites of vascular injury
The octapeptide pYMESRADR derived from integrin αIIb, previously shown to inhibit platelet aggregation by blocking fibrinogen binding to activated αIIbβ3 [27], corresponds to residues 313–320 of the insert loop (β-ribbon), that extends between the β2 and β3 strands of the W5 blade of the αIIb β-propeller domain [5]
Native pY313MESRADR320 was not able to inhibit αIIbβ31M Chinese hamster ovary (CHO) cell adhesion onto fibrinogen under shear stress. These results demonstrate that (i) the mutation of αIIb does not impact the binding of the peptide to the integrin in contrast to the β3K384A mutation and (ii) the residue pD319 in the octapeptide plays a crucial role in the binding of the octapeptide to integrin αIIbβ3

Summary

Introduction

Integrin αIIbβ, the platelet fibrinogen and von Willebrand factor (vWF) receptor, plays a key role in hemostasis and thrombosis, by promoting platelet aggregation and thrombus formation at sites of vascular injury. As circulating platelets in blood are constantly exposed to high. GlaxoSmithKline, Janssen, Lilly Canada, the Novartis Research Foundation, the Ontario Ministry of Economic Development and Innovation, Pfizer, Takeda, and the Wellcome Trust. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

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