Gain-of-Function Variant p.Pro2555Arg of von Willebrand Factor Increases Aggregate Size through Altering Stem Dynamics

Volker Huck; Po-Chia Chen; Emma-Ruoqi Xu; Alexander Tischer; Ulrike Klemm; Camilo Aponte-Santamaría; Christian Mess; Tobias Obser; Fabian Kutzki; Gesa König; Cécile V. Denis; Frauke Gräter; Matthias Wilmanns; Matthew Auton; Stefan W. Schneider; Reinhard Schneppenheim; Janosch Hennig; Maria A. Brehm

doi:10.1055/a-1344-4405

Thrombosis and Haemostasis, Inhaltsverzeichnis

CC BY-NC-ND 4.0 · Thromb Haemost 2022; 122(02): 226-239
DOI: 10.1055/a-1344-4405

Cellular Haemostasis and Platelets

Gain-of-Function Variant p.Pro2555Arg of von Willebrand Factor Increases Aggregate Size through Altering Stem Dynamics

Volker Huck‡^*

¹Department of Dermatology and Venereology, Center for Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

²Experimental Dermatology, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany

,

Po-Chia Chen‡^*

³Structural and Computational Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany

,

Emma-Ruoqi Xu‡^*

⁴European Molecular Biology Laboratory, Hamburg Unit, Hamburg, Germany

,

Alexander Tischer

⁵Division of Hematology, Mayo Clinic, Rochester, Minnesota, United States

,

Ulrike Klemm

⁶Department of Pediatric Hematology and Oncology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

,

Camilo Aponte-Santamaría

⁷Max Planck Tandem Group in Computational Biophysics, University of los Andes, Bogotá, Colombia

⁸Interdisciplinary Center for Scientific Computing, Heidelberg University, Heidelberg, Germany

,

Christian Mess

¹Department of Dermatology and Venereology, Center for Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

,

Tobias Obser

¹Department of Dermatology and Venereology, Center for Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

,

Fabian Kutzki

⁹Molecular Biomechanics Group, Heidelberg Institute for Theoretical Studies, Heidelberg, Germany

¹⁰Institute of Physical Chemistry, Karlsruhe Institute of Technology, Karlsruhe, Germany

,

Gesa König

⁶Department of Pediatric Hematology and Oncology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

,

Cécile V. Denis

¹¹Laboratory of Hemostasis, Inflammation and Thrombosis, Institut National de la Santé et de la Recherche Médicale UMR_1176, Université Paris-Saclay, Le Kremlin-Bicêtre, France

,

Frauke Gräter

⁸Interdisciplinary Center for Scientific Computing, Heidelberg University, Heidelberg, Germany

⁹Molecular Biomechanics Group, Heidelberg Institute for Theoretical Studies, Heidelberg, Germany

,

Matthias Wilmanns

⁴European Molecular Biology Laboratory, Hamburg Unit, Hamburg, Germany

¹²University Medical Center Hamburg-Eppendorf, Hamburg, Germany

,

Matthew Auton

⁵Division of Hematology, Mayo Clinic, Rochester, Minnesota, United States

,

Stefan W. Schneider

¹Department of Dermatology and Venereology, Center for Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

,

Reinhard Schneppenheim

⁶Department of Pediatric Hematology and Oncology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

,

Janosch Hennig^**

³Structural and Computational Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany

,

Maria A. Brehm^**

¹Department of Dermatology and Venereology, Center for Internal Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

› Institutsangaben

Abstract

The multimeric plasma glycoprotein (GP) von Willebrand factor (VWF) is best known for recruiting platelets to sites of injury during primary hemostasis. Generally, mutations in the VWF gene lead to loss of hemostatic activity and thus the bleeding disorder von Willebrand disease. By employing cone and platelet aggregometry and microfluidic assays, we uncovered a platelet GPIIb/IIIa-dependent prothrombotic gain of function (GOF) for variant p.Pro2555Arg, located in the C4 domain, leading to an increase in platelet aggregate size. We performed complementary biophysical and structural investigations using circular dichroism spectra, small-angle X-ray scattering, nuclear magnetic resonance spectroscopy, molecular dynamics simulations on the single C4 domain, and dimeric wild-type and p.Pro2555Arg constructs. C4-p.Pro2555Arg retained the overall structural conformation with minor populations of alternative conformations exhibiting increased hinge flexibility and slow conformational exchange. The dimeric protein becomes disordered and more flexible. Our data suggest that the GOF does not affect the binding affinity of the C4 domain for GPIIb/IIIa. Instead, the increased VWF dimer flexibility enhances temporal accessibility of platelet-binding sites. Using an interdisciplinary approach, we revealed that p.Pro2555Arg is the first VWF variant, which increases platelet aggregate size and shows a shear-dependent function of the VWF stem region, which can become hyperactive through mutations. Prothrombotic GOF variants of VWF are a novel concept of a VWF-associated pathomechanism of thromboembolic events, which is of general interest to vascular health but not yet considered in diagnostics. Thus, awareness should be raised for the risk they pose. Furthermore, our data implicate the C4 domain as a novel antithrombotic drug target.

Keywords

von Willebrand factor - hypercoagulability - GPIIb/IIIa

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Referenzen

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