Identification of new type 2B von Willebrand disease mutations: Arg543Gln,Arg545Pro and Arg578Leu

We report the identification in fire patients (three families) affected wit h type 2B ron Willebrand disease (VWD) of three heterozygous nucleotide sub stitutions at the codon for arginine 543, 545 and 578 of the mature non Wil lebrand factor (VWF) subunit resulting in a glutamine, proline and leucine substitution, respectively. These mutations are located in the Al loop wher e prevalent type 2B mutations (Arg543Trp, Arg545Cys and Arg578Gln) have bee n already identified at the same positions. By in vitro mutagenesis of full -length cDNA of VWF and transient expression in Cos-7 cells, we have shown that the six corresponding mutated recombinant VWFs (Gln543, Trp543, Cys545 , Pro545, Leu578 and Gln578 rVWF) exhibited quantitatively normal expressio n and normal multimeric pattern but increased ristocetin- and botrocetin-in duced binding to platelets as compared with that fur wildtype rVWF. The two mutations at position 545 induced the greatest reactivity for GPIb of corr esponding rVWFs as compared to the two mutations at positions 543 and 578.