| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1995: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1994: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1993: ¥700,000 (Direct Cost: ¥700,000)
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| Research Abstract |
The interaction of von Willebrand factor (vWF) and its one of the platelet receptorsglycoprotein (GP) Ib/IX complex, triggers platelet aggregation and plays a key role in regulating the initial step of platelet thrombus formation. In order tocharacterize the mechanisms that regualate the interaction, we have expressed a recombinant GP Ib/IX protein in mammalian cells and investigated the effect of amino acid substitutions or the changes in post-translational modifications on the receptor function. (1) We have analyzed the genomic DNA of patients with platelet-type von Willebrand disease, which is characterized by an abberant GPIb/IX with abnormally high affinity for vWF,and have found a point mutation in the alpha chain of the receptor. Furthemory, we have expressed in CHO cells the mutant pretein, which possessed the phenotypic abnormality of the disorder, the enhenced interaction with the ligand. (2) Substitutiuon of potentially sulfated three tyrosine residues with phenylalanine residues resulted in the loss of vWF binding function of the rteceptor. Recombinant protein expressed in CHO cells cultured in a condition that blocks protein tyrosine sulfation, i. e., in the presence of sodium chrolate, had impaired vWF binding function. (3) We have also improved the cell culture technique to obtain large amount of the recombinant protein, since this soluble recombinant protein inhibits the normal vWF binding to platelets thus having the potential role in the prevention of platelet thrombus formation. Our results provides essential informations for the basic mechanisms of arterial thrombosis and for the development of newantiplatelet therapies.
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