Simultaneous Exposure of Sites in von Willebrand Factor for Glycoprotein Ib Binding and ADAMTS13 Cleavage
Studies With Ristocetin
Objective—Platelet-bound von Willebrand factor (VWF) was recently demonstrated to be a better substrate for ADAMTS13, suggesting that 1 conformational change exposes both the glycoprotein Ibα binding site in the A1 domain and the ADAMTS13 cleavage site in the A2 domain. Because ristocetin induces VWF to bind glycoprotein Ibα in the absence of shear stress, we evaluated whether it could also enhance ADAMTS13 proteolysis of VWF.
Methods and Results—We used several VWF sources: plasma, purified plasma VWF, recombinant VWF fragments encompassing A1A2A3, A1A2, and 2 A2 domains, 1 containing a ristocetin-binding site (Asp1459–His1472) and the other lacking it. Ristocetin accelerated ADAMTS13 cleavage of multimeric VWF and of each of the recombinant VWF fragments except for the A2 domain lacking the ristocetin-binding site. We also examined the effect of ristocetin on the conformation of the A2 domain by assessing its effect on the susceptibility of Met1606 at the ADAMTS13 cleavage site to be oxidized by hypochlorous acid. Ristocetin markedly enhanced oxidation of Met1606 and Met1521 of the A2 domain.
Conclusion—These data indicate that exposure of the sites for glycoprotein Ibα and ADAMTS13 are coupled, explaining why platelet-bound VWF is a better ADAMTS13 substrate and why enhanced proteolysis is often observed in type 2B von Willebrand disease.
- Received May 29, 2012.
- Accepted August 6, 2012.
- © 2012 American Heart Association, Inc.