Molecular Modeling of Ligand and Mutation Sites of the Type A Domains of Human von Willebrand Factor and Their Relevance to von Willebrand's Disease

Abstract

Abstractvon Willebrand factor (vWF) is a large multimeric, multidomain glycoprotein found in platelets, endothelial cells and plasma. The A1, A2, and A3 domains in vWF mediate binding to glycoprotein Ib, ristocetin, botrocetin, collagen, sulphatides, and heparin and provide a protease cleavage site. Mutations causing types 2B, 2M, and 2A von Willebrand's disease (vWD) are located in the A1 and A2 domains. Homology modeling was performed to provide a molecular interpretation of vWF function and mutation sites. This was based on our previous alignment of 75 vWF-A sequences, the doubly wound α/β fold seen in recent vWF-A crystal structures from complement receptor type 3 and lymphocyte function-associated antigen-1, and our new alignment of 28 vWF A1 and A2 sequences from different species. The active site in doubly-wound α/β folds forms a crevice that is located at the switch point between the two halves of the central β-sheet, and usually contains two metal-binding Asp residues in the vWF-A superfamily. Although one of these Asp residues is absent from the A1, A2, and A3 domains, this crevice is shown to correspond to the ristocetin binding site in the A1 domain and the protease cleavage site in the A2 domain. The residues R571-K572-R578-R579-K585 are found to be conserved in 28 A1 sequences and are predicted to constitute the heparin binding site in the A1 domain. Inspection of the type 2M vWD mutation sites that are involved in downregulation of glycoprotein Ib (GpIb) binding to vWF shows that these are spatially clustered at the carboxyl-edge of the β-sheet and above it in the A1 domain and may directly perturb GpIb binding. In contrast, the type 2B vWD mutation sites that are involved in upregulation of GpIb binding to vWF are spatially clustered at the amino edge of this β-sheet and below it and are located on the opposite side of the A1 domain from the type 2M mutation sites. The type 2B mutations are located between the heparin and GpIb binding sites. Because heparin binding inhibits the interaction with GpIb, this provides an explanation of vWF upregulation. The type 2A vWD mutation sites in the A2 domain correspond to buried residues that are otherwise 100% conserved across all 28 species, and are likely to be important for the correct folding of the A2 domain and its physiologically important protease site.

FRETS‐VWF73, a first fluorogenic substrate for ADAMTS13 assay

“…The Y1605–M1606 bond is inaccessible in native VWF and made sensitive to ADAMTS13 by denaturation and shear force. Structural modelling has suggested that the bond is buried in the core β ‐sheet of the VWF A2 domain (Jenkins et al , 1998; Sutherland et al , 2004). This partially explains the requirement for denaturants or shear force in the hydrolysis of the Y1605–M1606 bond by ADAMTS13.…”

Type 2A (IIH) von Willebrand disease is due to mutations that affect von Willebrand factor multimerization

“…None of our in vitro experiments supported the hypothesis that the R1583Q substitution might be responsible for the propositus phenotype. The finding that, in the VWFs of many mammals, a glutamine is present at codon 1583 minimizes the role of this amino acid substitution [29]. In contrast, the R202W and C849Y mutations induce drastic amino acid changes, because in the former the small positively charged arginine is replaced by the bulk uncharged tryptophan, and in the latter the loss of a cysteine destroys a disulfide bond [30].…”

ABO blood group determines plasma von Willebrand factor levels: a biologic function after all?

“…60 Moreover, alignment of the VWF A2 domain sequences across 28 different mammalian species has shown that the consensus sequences for N-linked glycosylation are conserved at both asparagine 1515 and 1574. 61 Previous studies have demonstrated that N-linked glycan structures can directly influence the folding of glycoproteins, by reducing conformational freedom of the local peptide backbone. 62 To further test this hypothesis, we collected plasma from a series of 47 individuals with the rare Bombay phenotype.…”