Phenotype changes resulting in high-affinity binding of von Willebrand factor to recombinant glycoprotein Ib-IX: analysis of the platelet-type von Willebrand disease mutations

To maintain hemostasis under shear conditions, there must be an interaction between the platelet glycoprotein (GP) Ib-IX receptor and the plasma ligan d von Willebrand factor (vWf). In platelet-type von Willebrand disease (Pt- vWD), hemostasis is compromised. Two mutations in the GPIb alpha polypeptid e chain have been identified in these patients-a glycine-233 to valine chan ge and a methionine-239 to valine change. For this investigation, these mut ant proteins have been expressed in a Chinese hamster ovary cell model syst em. Ligand-binding studies were performed at various concentrations of rist ocetin, and adhesion assays were performed under flow conditions. The Pt-vW D mutations resulted in a gain-of-function receptor. vWf binding was increa sed at all concentrations of ristocetin examined, and adhesion on a vWf mat rix was enhanced in terms of cell tethering, slower rolling velocity, and d ecreased detachment with increasing shear rate. Two other mutations were al so introduced into the GPIb alpha chain. One mutation, encompassing both th e Pt-vWD mutations, created an increase in the hydrophobicity of this regio n. The second mutation, involving a valine-234 to glycine change, decreased the hydrophobicity of this region. Both mutations also resulted in a gain- of-function receptor, with the double mutation producing a hyperreactive re ceptor for vWf. These data further support the hypothesis that ligand bindi ng is regulated by conformational changes in the amino-terminal region of G PIb alpha, thereby influencing the stability of the GPIb alpha -vWf interac tion. (C) 2001 by The American Society of Hematology.