@article{3e6329ea63724ae79c5ef5526ab728c9,
title = "Evidence for the Misfolding of the A1 Domain within Multimeric von Willebrand Factor in Type 2 von Willebrand Disease",
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.",
keywords = "Hydrogen-deuterium exchange mass spectrometry, Limited proteolysis, Local disorder, Protein misfolding, von Willebrand factor",
author = "Alexander Tischer and Brehm, {Maria A.} and Machha, {Venkata R.} and Laurie Moon-Tasson and Benson, {Linda M.} and Nelton, {Katelynn J.} and Leger, {Rachel R.} and Tobias Obser and Marina Martinez-Vargas and Whitten, {Steven T.} and Dong Chen and Pruthi, {Rajiv K.} and Bergen, {H. Robert} and Cruz, {Miguel A.} and Reinhard Schneppenheim and Matthew Auton",
note = "Funding Information: This study was supported, in part, by research funding from the National Institutes of Health Grant HL109109 from NHLBI (M.A.), the Mayo Clinic Division of Hematology Small Grants Program ( CCaTS UL1TR000135 M.A.), the Mayo Clinic Department of Laboratory Medicine and Pathology Collaborative Research Funds (M.A., D.C.), the Mayo Clinic Center for Biomedical Discovery (M.A.), the Great Lakes Hemophilia Foundation and Health Resources and Services Administration through the Mayo Clinic Comprehensive Hemophilia Treatment Center (M.A., R.K.P.), and the German Research Foundation (DFG) to the Research Group FOR1543: Shear flow regulation of hemostasis – bridging the gap between nanomechanics and clinical presentation (R.S., M.A.B., T.O.). The authors thank the technical support from staff of the Mayo Clinic Special Coagulation Laboratory , the Mayo Clinic Proteomics Core for technical support with limited trypsinolysis mass spectrometry and analysis, and the UKE Microscopy Imaging Facility (UMIF) for technical support and providing the Leica SP5 microscope. The authors also gratefully acknowledge Drs. S. Walter Englander and Leland Mayne for very helpful scientific discussions regarding optimization of HXMS and the establishment of HXMS technology in their laboratory. In addition, the authors also acknowledge charitable contributions from Mark Davies' Cycle von Willebrand Disease, which have defrayed, in part, publication costs. Funding Information: This study was supported, in part, by research funding from the National Institutes of Health Grant HL109109 from NHLBI (M.A.), the Mayo Clinic Division of Hematology Small Grants Program (CCaTS UL1TR000135 M.A.), the Mayo Clinic Department of Laboratory Medicine and Pathology Collaborative Research Funds (M.A. D.C.), the Mayo Clinic Center for Biomedical Discovery (M.A.), the Great Lakes Hemophilia Foundation and Health Resources and Services Administration through the Mayo Clinic Comprehensive Hemophilia Treatment Center (M.A. R.K.P.), and the German Research Foundation (DFG) to the Research Group FOR1543: Shear flow regulation of hemostasis ? bridging the gap between nanomechanics and clinical presentation (R.S. M.A.B. T.O.). The authors thank the technical support from staff of the Mayo Clinic Special Coagulation Laboratory, the Mayo Clinic Proteomics Core for technical support with limited trypsinolysis mass spectrometry and analysis, and the UKE Microscopy Imaging Facility (UMIF) for technical support and providing the Leica SP5 microscope. The authors also gratefully acknowledge Drs. S. Walter Englander and Leland Mayne for very helpful scientific discussions regarding optimization of HXMS and the establishment of HXMS technology in their laboratory. In addition, the authors also acknowledge charitable contributions from Mark Davies' Cycle von Willebrand Disease, which have defrayed, in part, publication costs. Publisher Copyright: {\textcopyright} 2019 Elsevier Ltd",
year = "2020",
month = jan,
day = "17",
doi = "10.1016/j.jmb.2019.09.022",
language = "English (US)",
volume = "432",
pages = "305--323",
journal = "Journal of Molecular Biology",
issn = "0022-2836",
publisher = "Academic Press Inc.",
number = "2",
}