Type 2M von Willebrand Disease: F606I and I662F Mutations in the Glycoprotein Ib Binding Domain Selectively Impair Ristocetin- but not Botrocetin-Mediated Binding of von Willebrand Factor to Platelets

Abstract

Abstractvon Willebrand disease (vWD) is a common, autosomally inherited, bleeding disorder caused by quantitative and/or qualitative deficiency of von Willebrand factor (vWF). We describe two families with a variant form of vWD where affected members of both families have borderline or low vWF antigen levels, normal vWF multimer patterns, disproportionately low ristocetin cofactor activity, and significant bleeding symptoms. Whereas ristocetin-induced binding of plasma vWF from affected members of both families to fixed platelets was reduced, botrocetin-induced platelet binding was normal. The sequencing of genomic DNA identified unique missense mutations in each family in the vWF exon 28. In Family A, a missense mutation at nucleotide 4105T → A resulted in a Phe606Ile amino acid substitution (F606I) and in Family B, a missense mutation at nucleotide 4273A → T resulted in an Ile662Phe amino acid substitution (I662F). Both mutations are within the large disulfide loop between Cys509 and Cys695 in the A1 domain that mediates vWF interaction with platelet glycoprotein Ib. Expression of recombinant vWF containing either F606I or I662F mutations resulted in mutant recombinant vWF with decreased ristocetin-induced platelet binding, but normal multimer structure, botrocetin-induced platelet binding, collagen binding, and binding to the conformation-sensitive monoclonal antibody, AvW-3. Both mutations are phenotypically distinct from the previously reported variant type 2MMilwaukee-1 because of the presence of normal botrocetin-induced platelet binding, collagen binding, and AvW-3 binding, as well as the greater frequency and intensity of clinical bleeding. When the reported type 2M mutations are mapped on the predicted three-dimensional structure of the A1 loop of vWF, the mutations cluster in one region that is distinct from the region in which the type 2B mutations cluster.

N1421K mutation in the glycoprotein Ib binding domain impairs ristocetin‐ and botrocetin‐mediated binding of von Willebrand factor to platelets

“…This variant is characterized by the full range of VWF multimers in plasma and decreased VWFdependent platelet adhesion, usually revealed as low VWF:RCo (1). A somewhat reduced quantity of VWF:Ag has also occasionally been observed in other patients with type 2M (11,22). Type 2M mutations are clustered in the A1 domain containing the GPIb binding site.…”

“…Type 2M mutations are clustered in the A1 domain containing the GPIb binding site. To date, 17 mutations causing type 2M VWD have been identified (11,(22)(23)(24)(25)(26)(27)(28)(29).…”

“…Amino acid substitutions leading to gain-of-function typical of type 2B are clustered in a section of the A1 domain opposite to the GPIb binding site and believed to relieve a normal inhibitory effect of this region (30,31). Mutations that cause type 2M (11,(23)(24)(25) are often distributed around Lys 1362 and may prevent the correct exposure of the GPIb binding site (32). The N1421K mutation impaired ristocetin as well as botrocetin-induced VWF binding to GPIb.…”

Charcot–Marie–Tooth Disease type 4C: Novel mutations, clinical presentations, and diagnostic challenges

“…Individual diagnosis of CMT4C patients is challenging because of considerable intrafamilial and interfamilial clinical variability. The diagnostic challenge was further compounded in sporadic CMT4C patients because of the presence of additional extremely rare inherited disorders such as SCADD, 16 retinitis pigmentosa, 17 von Willebrand type 2M, 18 and PAI-1 deficiency. 19 When considering our patients together with previous reports from the literature, it seems appropriate for CMT4C to be considered an atypical form of CMT.…”

The molecular biology of von Willebrand disease

“…Interestingly, amino acid 1324 lies within the main cluster of type 2B VWD mutations (amino acids 1303±1341). The candidate missense mutations, F1369I (Phe606Ile) (Gaucher et al, 1995a) and I1425P (Ile662Phe) (Hillery et al, 1998), have been con®rmed in two separate families with a type 2M phenotype. Furthermore, molecular modelling of the three dimensional structure of the VWF A1 domain predicts that these mutations are not in the same cluster as the majority of type 2B VWD mutations (Hillery et al, 1998;Jenkins et al, 1998).…”