Mechanical properties of VWF in single molecule and cell adhesion force experiments using AFM
Zusammenfassung der Projektergebnisse
Excessive blood loss at a site of vascular injury is prevented by recruitment of platelets to the injured vessel wall and the formation of a platelet plug. These processes are critically mediated by the plasma glycoprotein von Willebrand factor (VWF), which circulates in the blood in the form of long, linear multimers. Remarkably, VWF’s activation for hemostasis is force-induced: When subjected to elevated hydrodynamic forces in the blood flow, which primarily occur in connection with blood vessel injury, VWF undergoes several conformational changes – both on the level of single domains and of the overall protein conformation – that lead to increased affinity for binding collagen and platelets. Furthermore, also VWF’s down-regulation is force-regulated, as it relies on enzymatic cleavage of a cleavage site that is accessible only upon force-induced unfolding of VWF’s A2 domains. In order to elucidate the molecular mechanisms that underlie VWF’s extraordinary response to force and the associated conformational changes, we combined atomic force microscope (AFM)-based single-molecule force spectroscopy and AFM imaging to directly probe the mechanics and structure of VWF dimers under a variety of different conditions. Key results were the discovery of a strong intermonomer interaction in VWF mediated by its D4 domains that is expected to tune VWF’s ability to sense hydrodynamic forces in the bloodstream, and the finding that even minute pH changes markedly affect VWF’s conformation and its response to force. In addition, our combined approach indicated the existence of further physiologically relevant interactions of the C-domains within VWF dimers that, however, dissociate at forces too low to be resolved directly by the AFM. Therefore, we currently employ magnetic tweezers-based singlemolecule force spectroscopy to probe VWF’s behavior at very low forces of ≈1 pN, at which first conformational transitions of VWF multimers are thought to occur. Taken together, our data refined our picture of VWF’s conformational transitions under elevated hydrodynamic forces. They will thus help to comprehend the force-induced activation of VWF and provide clues for understanding clotting disorders, such as von Willebrand disease and thrombosis, at the single-molecule level.
Projektbezogene Publikationen (Auswahl)
- VWF–Collagen Interactions Studied with Single Molecule Force Spectroscopy. Biophys J. 2014;106 (2):450a
Posch S, Obser T, Brehm AM, Gruber HJ, Schneppenheim R, Tampé R, Hinterdorfer P
(Siehe online unter https://doi.org/10.1016/j.bpj.2013.11.2551) - Collagen Surface Functionalization - A New Strategy for Molecular Recognition Force Spectroscopy. Imaging & Microscopy 2015 Mar(1)
Posch S
- Force-sensitive autoinhibition of the von Willebrand factor is mediated by interdomain interactions. Biophys J. 2015 May 5;108(9):2312-21
Aponte-Santamaría C, Huck V, Posch S, Bronowska AK, Grässle S, Brehm MA, Obser T, Schneppenheim R, Hinterdorfer P, Schneider SW, Baldauf C, Gräter F
(Siehe online unter https://doi.org/10.1016/j.bpj.2015.03.041) - Mechanosensitive Von Willebrand Factor Protein-Protein Interactions Regulate Hemostasis. Biophys J. 2015; 108 (2): 505a
Aponte-Santamaría C, Huck V, Posch S, Bronowska AK, Grässle S, Brehm AM, Obser T, Schneppenheim R, Hinterdorfer P, Schneider SW, Baldauf C, Gräter F
(Siehe online unter https://doi.org/10.1016/j.bpj.2014.11.2764) - Single molecule force spectroscopy data and BD- and MD simulations on the blood protein von Willebrand factor. Data Brief. 2016 Jul 21;8:1080-7
Posch S, Aponte-Santamaría C, Schwarzl R, Karner A, Radtke M, Gräter F, Obser T, König G, Brehm MA, Gruber HJ, Netz RR, Baldauf C, Schneppenheim R, Tampé R, Hinterdorfer P
(Siehe online unter https://doi.org/10.1016/j.dib.2016.07.031) - Mutual A domain interactions in the force sensing protein von Willebrand factor. J Struct Biol. 2017 Jan;197(1):57-64
Posch S, Aponte-Santamaría C, Schwarzl R, Karner A, Radtke M, Gräter F, Obser T, König G, Brehm MA, Gruber HJ, Netz RR, Baldauf C, Schneppenheim R, Tampé R, Hinterdorfer P
(Siehe online unter https://doi.org/10.1016/j.jsb.2016.04.012) - Interaction of von Willebrand factor domains with collagen investigated by single molecule force spectroscopy. J Chem Phys. 2018 Mar 28;148(12):123310
Posch S, Obser T, König G, Schneppenheim R, Tampé R, Hinterdorfer P
(Siehe online unter https://doi.org/10.1063/1.5007313)
