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  Von Willebrand factor is dimerized by protein disulfide isomerase

Lippok, S., Kolšek, K., Löf, A., Eggert, D., Vanderlinden, W., Müller, J. P., et al. (2016). Von Willebrand factor is dimerized by protein disulfide isomerase. Blood, 127(9), 1183-1191. doi:10.1182/blood-2015-04-641902.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-002A-1719-1 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-002A-171A-0
Genre: Journal Article

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http://dx.doi.org/10.1182/blood-2015-04-641902 (Publisher version)
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 Creators:
Lippok, Svenja1, Author
Kolšek, Katra2, Author
Löf, Achim1, Author
Eggert, Dennis3, 4, Author              
Vanderlinden, Willem1, 5, Author
Müller, Jochen P.1, Author
König, Gesa6, Author
Obser, Tobias6, Author
Röhrs, Karoline6, Author
Schneppenheim, Sonja7, Author
Budde, Ulrich7, Author
Baldauf, Carsten8, Author              
Aponte-Santamaría, Camilo2, Author
Gräter, Frauke2, Author
Schneppenheim, Reinhard6, Author
Rädler, Joachim O.1, Author
Brehm, Maria A.6, Author
Affiliations:
1Faculty of Physics and Center for NanoScience, Ludwig Maximilian University, Munich, Germany, ou_persistent22              
2Molecular Biomechanics Group, Heidelberg Institute for Theoretical Studies, Heidelberg, Germany, ou_persistent22              
3Miller Group, Atomically Resolved Dynamics Department, Max Planck Institute for the Structure and Dynamics of Matter, Max Planck Society, ou_1938288              
4Microscopy and Image Analysis Technology Platform, Heinrich-Pette-Institute, Leibniz Institute for Experimental Virology, Hamburg, Germany, ou_persistent22              
5Department of Chemistry, Division of Molecular Imaging and Photonics, Katholieke Universiteit Leuven-University of Leuven, Leuven, Belgium, ou_persistent22              
6Department of Pediatric Hematology and Oncology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany, ou_persistent22              
7Medilys Laborgesellschaft mbH, Hemostaseology, Asklepios Klinik Altona, Hamburg, Germany, ou_persistent22              
8Theory, Fritz Haber Institute, Max Planck Society, ou_634547              

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 Abstract: Multimeric von Willebrand factor (VWF) is essential for primary hemostasis. The biosynthesis of VWF high-molecular-weight multimers requires spatial separation of each step because of varying pH value requirements. VWF is dimerized in the endoplasmic reticulum by formation of disulfide bonds between the C-terminal cysteine knot (CK) domains of 2 monomers. Here, we investigated the basic question of which protein catalyzes the dimerization. We examined the putative interaction of VWF and the protein disulfide isomerase PDIA1, which has previously been used to visualize endoplasmic reticulum localization of VWF. Excitingly, we were able to visualize the PDI–VWF dimer complex by high-resolution stochastic optical reconstruction microscopy and atomic force microscopy. We proved and quantified direct binding of PDIA1 to VWF, using microscale thermophoresis and fluorescence correlation spectroscopy (dissociation constants KD = 236 ± 66 nM and KD = 282 ± 123 nM by microscale thermophoresis and fluorescence correlation spectroscopy, respectively). The similar KD (258 ± 104 nM) measured for PDI interaction with the isolated CK domain and the atomic force microscopy images strongly indicate that PDIA1 binds exclusively to the CK domain, suggesting a key role of PDIA1 in VWF dimerization. On the basis of protein–protein docking and molecular dynamics simulations, combined with fluorescence microscopy studies of VWF CK-domain mutants, we suggest the following mechanism of VWF dimerization: PDI initiates VWF dimerization by forming the first 2 disulfide bonds Cys2771-2773′ and Cys2771′-2773. Subsequently, the third bond, Cys2811-2811′, is formed, presumably to protect the first 2 bonds from reduction, thereby rendering dimerization irreversible. This study deepens our understanding of the mechanism of VWF dimerization and the pathophysiological consequences of its inhibition.

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Language(s): eng - English
 Dates: 2015-04-222015-12-082015-12-152016-03-03
 Publication Status: Published in print
 Pages: 9
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: DOI: 10.1182/blood-2015-04-641902
 Degree: -

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Title: Blood
  Other : Blood
Source Genre: Journal
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Publ. Info: Philadelphia, Pa. : W.B. Saunders
Pages: - Volume / Issue: 127 (9) Sequence Number: - Start / End Page: 1183 - 1191 Identifier: ISSN: 0006-4971
CoNE: https://pure.mpg.de/cone/journals/resource/954925385125