title: Force-sensitive autoinhibition of the von Willebrand factor is mediated by interdomain interactions.
abstract: Von Willebrand factor (VWF) plays a central role in hemostasis. Triggered by shear-stress, it adheres to platelets at sites of vascular injury. Inactivation of VWF has been associated to the shielding of its adhesion sites and proteolytic cleavage. However, the molecular nature of this shielding and its coupling to cleavage under shear-forces in flowing blood remain unknown. In this study, we describe, to our knowledge, a new force-sensory mechanism for VWF-platelet binding, which addresses these questions, based on a combination of molecular dynamics (MD) simulations, atomic force microscopy (AFM), and microfluidic experiments. Our MD simulations demonstrate that the VWF A2 domain targets a specific region at the VWF A1 domain, corresponding to the binding site of the platelet glycoprotein Ibα (GPIbα) receptor, thereby causing its blockage. This implies autoinhibition of the VWF for the binding of platelets mediated by the A1-A2 protein-protein interaction. During force-probe MD simulations, a stretching force dissociated the A1A2 complex, thereby unblocking the GPIbα binding site. Dissociation was found to be coupled to the unfolding of the A2 domain, with dissociation predominantly occurring before exposure of the cleavage site in A2, an observation that is supported by our AFM experiments. This suggests that the A2 domain prevents platelet binding in a force-dependent manner, ensuring that VWF initiates hemostasis before inactivation by proteolytic cleavage. Microfluidic experiments with an A2-deletion VWF mutant resulted in increased platelet binding, corroborating the key autoinhibitory role of the A2 domain within VWF multimers. Overall, autoinhibition of VWF mediated by force-dependent interdomain interactions offers the molecular basis for the shear-sensitive growth of VWF-platelet aggregates, and might be similarly involved in shear-induced VWF self-aggregation and other force-sensing functions in hemostasis.
authors.affiliation: Molecular Biomechanics Group, Heidelberg Institute for Theoretical Studies, Heidelberg, Germany.
authors.forename: Camilo
authors.initials: C
authors.lastname: Aponte-Santamaría
authors.affiliation: Experimental Dermatology, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany.
authors.forename: Volker
authors.initials: V
authors.lastname: Huck
authors.affiliation: Department of Applied Experimental Biophysics, Institute of Biophysics, Johannes Kepler University, Linz, Austria.
authors.forename: Sandra
authors.initials: S
authors.lastname: Posch
authors.affiliation: Molecular Biomechanics Group, Heidelberg Institute for Theoretical Studies, Heidelberg, Germany.
authors.forename: Agnieszka K
authors.initials: AK
authors.lastname: Bronowska
authors.affiliation: Experimental Dermatology, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany.
authors.forename: Sandra
authors.initials: S
authors.lastname: Grässle
authors.affiliation: Department of Pediatric Hematology and Oncology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: Maria A
authors.initials: MA
authors.lastname: Brehm
authors.affiliation: Department of Pediatric Hematology and Oncology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: Tobias
authors.initials: T
authors.lastname: Obser
authors.affiliation: Department of Pediatric Hematology and Oncology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Schneppenheim
authors.affiliation: Department of Applied Experimental Biophysics, Institute of Biophysics, Johannes Kepler University, Linz, Austria.
authors.forename: Peter
authors.initials: P
authors.lastname: Hinterdorfer
authors.affiliation: Experimental Dermatology, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany.
authors.forename: Stefan W
authors.initials: SW
authors.lastname: Schneider
authors.affiliation: Theory Department, Fritz-Haber-Institut der Max-Planck-Gesellschaft, Berlin, Germany. Electronic address: baldauf@fhi-berlin.mpg.de.
authors.forename: Carsten
authors.initials: C
authors.lastname: Baldauf
authors.affiliation: Molecular Biomechanics Group, Heidelberg Institute for Theoretical Studies, Heidelberg, Germany. Electronic address: frauke.graeter@h-its.org.
authors.forename: Frauke
authors.initials: F
authors.lastname: Gräter
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D006801:Humans; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000378:metabolism; D014841:von Willebrand Factor / Q000737:chemistry* / Q000378:metabolism
chemical_list: D019038:Platelet Glycoprotein GPIb-IX Complex; C094990:adhesion receptor; D014841:von Willebrand Factor