Expression and functional characterization of an abnormal platelet membrane glycoprotein Ibα (Met²³⁹ → Val) reported in patients with platelet-type yon Willebrand Disease

Platelet-type von Willebrand disease (vWD) is a congenital bleeding disorder characterized by heightened ristocetin-induced platelet aggregation caused by abnormally high affinity between the platelet membrane glycoprotein (GP) Ib/IX complex and von Willebrand factor (vWF). Two distinct point mutations, Gly²³³ to Val and Met²³⁹ to Val, have been reported in GPIbα. We have constructed a recombinant GPIbα fragment containing the latter mutation, Met²³⁹ to Val (M239V) and characterized the mutant molecule using two methods, ie, interaction between soluble vWF and immobilized M239V and inhibition of platelet aggregation by purified soluble M239V. Spontaneous binding (ie, binding without any inducers) was observed between ¹²⁵I-vWF and immobilized M239V but not between ¹²⁵I-vWF and immobilized wild-type (WT) GPIbα. The addition of low concentrations of ristocetin (0.2 mg/mL) induced specific ¹²⁵I-vWF binding to immobilized M239V, but not to WT GPIbα. At high concentrations of ristocetin (1.2 mg/mL), both WT GPIbα and M239V specifically bound to ¹²⁵I-vWF. Thus, M239V reproduced the unique functional abnormality of the GPIb/IX complex in platelet-type vWD. Moreover, the purified soluble M239V inhibited platelet aggregation induced by low concentration of ristocetin (0.3 mg/mL) in platelet-rich plasma from a patient having Met²³⁸ to Val mutation, whereas purified WT did not. These results provide direct evidences that the reported point mutation is the responsible molecular basis of this disorder.