Abstract

Background Shear stress triggers conformational stretching of von Willebrand factor (VWF), responsible for its self-association and binding to the platelet receptor GpIbα. This phenomenon supports primary haemostasis under flow. Type 2B-VWF natural mutants are considered having increased affinity for platelet GpIbα. Objectives To assess the mechanism responsible for the enhanced interaction of the p.R1306W- VWF mutant with the platelet receptor. Methods The interaction of GpIbα with WT and p.R1306W-VWF multimers and A1-A2-A3 constructs was investigated with surface plasmon resonance spectroscopy. The analysis of static VWF conformation in solution was carried out by dynamic light scattering spectroscopy. The shear-stress induced self-association of VWF multimers was investigated by atomic force microscopy (AFM) over a 0-60 dyn/cm2 range. Results WT-VWF does not interact with GpIbα under static conditions, whereas the mutant at ≈2 μg/ml already binds to the receptor. By contrast, the WT- and p.R1306W-A1-A2-A3 constructs showed comparable affinities for GpIbα (Kd ≈ 20 nM). The hydrodynamic diameter of resting R1306W -VWF multimers was significantly higher than the WT (210 ± 60 nm vs. 87 ± 22 nm). At shear forces<14 dyn/cm2, the p.R1306W multimers rapidly change conformation entering a regime of self-aggregation, which, at variance, is induced for WT-VWF by shear forces>30 dyn/cm2. Mechanical stretching AFM experiments showed that p.R1306W multimers need less energy/length unit (≈ 10 pN) to be stretched compared to WT protein. Conclusions The increased avidity of p.R1306W-VWF for GpIbα arises mostly from a higher sensitivity to shear stress that facilitates exposure of GpIbα binding sites.
Lingua originaleEnglish
pagine (da-a)n/a-n/a
Numero di pagine7
RivistaJournal of Thrombosis and Haemostasis
Volume2013
DOI
Stato di pubblicazionePubblicato - 2013

Keywords

  • Atomic Force Microscopy
  • Shear stress
  • Surface Plasmon Resonance
  • Von Willebrand Disease

Fingerprint Entra nei temi di ricerca di 'The Type 2b P.R1306w Natural Mutation Of Von Willebrand Factor Dramatically Enhances The Multimer Sensitivity To Shear Stress'. Insieme formano una fingerprint unica.

Cita questo