Evidence for the Misfolding of the A1 Domain within Multimeric von Willebrand Factor in Type 2 von Willebrand Disease

Alexander Tischer, Maria A. Brehm, Venkata R. Machha, Laurie Moon-Tasson, Linda M. Benson, Katelynn J. Nelton, Rachel R. Leger, Tobias Obser, Marina Martinez-Vargas, Steven T. Whitten, Dong Chen, Rajiv K. Pruthi, H. Robert Bergen, Miguel A. Cruz, Reinhard Schneppenheim, Matthew Auton

Research output: Contribution to journalArticle

Abstract

Von Willebrand factor (VWF), an exceptionally large multimeric plasma glycoprotein, functions to initiate coagulation by agglutinating platelets in the blood stream to sites of vascular injury. This primary hemostatic function is perturbed in type 2 dysfunctional subtypes of von Willebrand disease (VWD) by mutations that alter the structure and function of the platelet GPIbα adhesive VWF A1 domains. The resulting amino acid substitutions cause local disorder and misfold the native structure of the isolated platelet GPIbα–adhesive A1 domain of VWF in both gain-of-function (type 2B) and loss-of-function (type 2M) phenotypes. These structural effects have not been explicitly observed in A1 domains of VWF multimers native to blood plasma. New mass spectrometry strategies are applied to resolve the structural effects of 2B and 2M mutations in VWF to verify the presence of A1 domain structural disorder in multimeric VWF harboring type 2 VWD mutations. Limited trypsinolysis mass spectrometry (LTMS) and hydrogen-deuterium exchange mass spectrometry (HXMS) are applied to wild-type and VWD variants of the single A1, A2, and A3 domains, an A1A2A3 tridomain fragment of VWF, plasmin-cleaved dimers of VWF, multimeric recombinant VWF, and normal VWF plasma concentrates. Comparatively, these methods show that mutations known to misfold the isolated A1 domain increase the rate of trypsinolysis and the extent of hydrogen-deuterium exchange in local secondary structures of A1 within multimeric VWF. VWD mutation effects are localized to the A1 domain without appreciably affecting the structure and dynamics of other VWF domains. The intrinsic dynamics of A1 observed in recombinant fragments of VWF are conserved in plasma-derived VWF. These studies reveal that structural disorder does occur in VWD variants of the A1 domain within multimeric VWF and provides strong support for VWF misfolding as a result of some, but not all, type 2 VWD variants.

Original languageEnglish (US)
Pages (from-to)305-323
Number of pages19
JournalJournal of Molecular Biology
Volume432
Issue number2
DOIs
StatePublished - Jan 17 2020

Keywords

  • Hydrogen-deuterium exchange mass spectrometry
  • Limited proteolysis
  • Local disorder
  • Protein misfolding
  • von Willebrand factor

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

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  • Cite this

    Tischer, A., Brehm, M. A., Machha, V. R., Moon-Tasson, L., Benson, L. M., Nelton, K. J., Leger, R. R., Obser, T., Martinez-Vargas, M., Whitten, S. T., Chen, D., Pruthi, R. K., Bergen, H. R., Cruz, M. A., Schneppenheim, R., & Auton, M. (2020). Evidence for the Misfolding of the A1 Domain within Multimeric von Willebrand Factor in Type 2 von Willebrand Disease. Journal of Molecular Biology, 432(2), 305-323. https://doi.org/10.1016/j.jmb.2019.09.022