title: Structural basis of type 2A von Willebrand disease investigated by molecular dynamics simulations and experiments.
abstract: The hemostatic function of von Willebrand factor is downregulated by the metalloprotease ADAMTS13, which cleaves at a unique site normally buried in the A2 domain. Exposure of the proteolytic site is induced in the wild-type by shear stress as von Willebrand factor circulates in blood. Mutations in the A2 domain, which increase its susceptibility to cleavage, cause type 2A von Willebrand disease. In this study, molecular dynamics simulations suggest that the A2 domain unfolds under tensile force progressively through a series of steps. The simulation results also indicated that three type 2A mutations in the C-terminal half of the A2 domain, L1657I, I1628T and E1638K, destabilize the native state fold of the protein. Furthermore, all three type 2A mutations lowered in silico the tensile force necessary to undock the C-terminal helix α6 from the rest of the A2 domain, the first event in the unfolding pathway. The mutations F1520A, I1651A and A1661G were also predicted by simulations to destabilize the A2 domain and facilitate exposure of the cleavage site. Recombinant A2 domain proteins were expressed and cleavage assays were performed with the wild-type and single-point mutants. All three type 2A and two of the three predicted mutations exhibited increased rate of cleavage by ADAMTS13. These results confirm that destabilization of the helix α6 in the A2 domain facilitates exposure of the cleavage site and increases the rate of cleavage by ADAMTS13.
authors.affiliation: Department of Bioengineering, University of Washington, Seattle, Washington, United States of America. gianluca@u.washington.edu
authors.forename: Gianluca
authors.initials: G
authors.lastname: Interlandi
authors.forename: Minhua
authors.initials: M
authors.lastname: Ling
authors.forename: An Yue
authors.initials: AY
authors.lastname: Tu
authors.forename: Dominic W
authors.initials: DW
authors.lastname: Chung
authors.forename: Wendy E
authors.initials: WE
authors.lastname: Thomas
mesh_terms: D051722:ADAM Proteins / Q000235:genetics / Q000378:metabolism; D000071120:ADAMTS13 Protein; D002460:Cell Line; D006801:Humans; D056004:Molecular Dynamics Simulation; D017433:Protein Structure, Secondary; D056728:von Willebrand Disease, Type 2 / Q000235:genetics / Q000378:metabolism*; D014841:von Willebrand Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D014841:von Willebrand Factor; D051722:ADAM Proteins; D000071120:ADAMTS13 Protein; C099604:ADAMTS13 protein, human