The interaction of von Willebrand factor-A1 domain with collagen: mutation G1324S (type 2M von Willebrand disease) impairs the conformational change in A1 domain induced by collagen.
BACKGROUND
It is established that the A3 domain in von Willebrand factor (VWF) contains the major collagen-binding site. However, there are conflicting reports describing the capacity of the A1 domain to interact with collagen types I and III.


METHODS
In this study, we have used recombinant VWF-A1 polypeptides, as well as conformation-specific monoclonal antibodies (mAb), to analyze the A1-collagen interaction.


RESULTS
The A1 domain bound to collagen with K(d) approximately 8.0 nm and this binding was blocked by the mAb 6G1, which blocks the interaction between ristocetin and VWF. In addition, collagen-bound A1 protein was able to support flow-dependent adhesion of platelets, demonstrating that the binding sites for collagen and glycoprotein (GP)Ib are different. Analysis with two conformation-specific mAb demonstrated that the structure of the A1 domain changed as a result of the binding to collagen. In contrast, the antibodies failed to detect conformational change in the G1324S mutant (type 2M von Willebrand disease). Thus, direct binding to collagen induces a change in the structural conformation within the VWF-A1 domain, and the G1324S substitution prevents this conformational change.


CONCLUSIONS
This study has shown that the isolated A1 domain can simultaneously bind to collagen and platelet GPIb, supporting platelet adhesion under high-flow conditions. In addition, this study has used mAb to demonstrate that the binding of the isolated A1 domain or full-length VWF to collagen is accompanied by a conformational change in A1 domain.
Morales
L D
LD
Department of Medicine, Thrombosis Research Section, Baylor College of Medicine, Houston, TX 77030, USA.
Martin
C
C
Cruz
M A
MA
D019943:Amino Acid Substitution; D000911:Antibodies, Monoclonal; D001665:Binding Sites; D003094:Collagen / Q000378:metabolism*; D024042:Collagen Type I / Q000378:metabolism; D024061:Collagen Type III / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008958:Models, Molecular; D017354:Point Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D020349:Surface Plasmon Resonance; D014842:von Willebrand Diseases / Q000097:blood* / Q000145:classification / Q000235:genetics*; D014841:von Willebrand Factor / Q000737:chemistry* / Q000235:genetics* / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D024042:Collagen Type I; D024061:Collagen Type III; D011994:Recombinant Proteins; D014841:von Willebrand Factor; D003094:Collagen