N-terminal flanking region of A1 domain in von Willebrand factor stabilizes structure of A1A2A3 complex and modulates platelet activation under shear stress

J Biol Chem. 2012 Apr 27;287(18):14579-85. doi: 10.1074/jbc.M112.348573. Epub 2012 Mar 19.

Abstract

von Willebrand factor (vWF) mediates platelet adhesion and thrombus formation via its interaction with the platelet receptor glycoprotein (GP)Ibα. We have analyzed two A1A2A3 tri-domain proteins to demonstrate that the amino acid sequence, Gln(1238)-Glu(1260), in the N-terminal flanking region of the A1 domain, together with the association between the A domains, modulates vWF-GPIbα binding and platelet activation under shear stress. Using circular dichroism spectroscopy and differential scanning calorimetry, we have described that sequence Gln(1238)-Glu(1260) stabilizes the structural conformation of the A1A2A3 tri-domain complex. The structural stabilization imparted by this particular region inhibits the binding capacity of the tri-domain protein for GPIbα. Deletion of this region causes a conformational change in the A1 domain that increases binding to GPIbα. Only the truncated protein was capable of effectively blocking ristocetin-induced platelet agglutination. To determine the capacity of activating platelets via the interaction with GPIbα, whole blood was incubated with the N-terminal region truncated or intact tri-A domain protein prior to perfusion over a fibrin(ogen)-coated surface. At a high shear rate of 1,500 s(-1), platelets from blood containing the truncated protein rapidly bound, covering >90% of the fibrin(ogen) surface area, whereas the intact tri-A domain protein induced platelets to bind <10%. The results obtained in this study ascertain the relevant role of the structural association between the N-terminal flanking region of the A1 domain (amino acids Gln(1238)-Glu(1260)) and the A1A2A3 domain complex in preventing vWF to bind spontaneously to GPIbα in solution under high shear forces.

Publication types

  • Clinical Trial
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anti-Bacterial Agents / pharmacology
  • Female
  • Fibrinogen / genetics
  • Fibrinogen / metabolism
  • Humans
  • Male
  • Membrane Glycoproteins / genetics
  • Membrane Glycoproteins / metabolism*
  • Multiprotein Complexes / genetics
  • Multiprotein Complexes / metabolism
  • Platelet Activation / drug effects
  • Platelet Activation / physiology*
  • Platelet Glycoprotein GPIb-IX Complex
  • Protein Structure, Quaternary
  • Protein Structure, Tertiary
  • Ristocetin / pharmacology
  • Stress, Physiological / drug effects
  • Stress, Physiological / physiology*
  • von Willebrand Factor / genetics
  • von Willebrand Factor / metabolism*

Substances

  • Anti-Bacterial Agents
  • Membrane Glycoproteins
  • Multiprotein Complexes
  • Platelet Glycoprotein GPIb-IX Complex
  • adhesion receptor
  • von Willebrand Factor
  • Ristocetin
  • Fibrinogen