TY - JOUR
T1 - A conformational transition of the D9D3 domain primes von Willebrand factor for multimerization
AU - Gruber, Sophia
AU - Lof, Achim
AU - Hausch, Adina
AU - Kutzki, Fabian
AU - Johr, Res
AU - Obser, Tobias
AU - Konig, Gesa
AU - Schneppenheim, Reinhard
AU - Aponte-Santamaría, Camilo
AU - Grater, Frauke
AU - Brehm, Maria A.
AU - Benoit, Martin
AU - Lipfert, Jan
N1 - Funding Information:
This project was funded by Deutsche Forschungsgemeinschaft (German Research Foundation) project 201269156 (SFB 1032), project 386143268 (“Unraveling the Mechano-Regulation of Von Willebrand Factor”), RTG/GRK 2450, and INST 40/467-1 FUGG, by the state of Baden-Wu€rttemberg through bwHPC, and by the Klaus Tschira Foundation.
Publisher Copyright:
© 2022 by The American Society of Hematology. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.
PY - 2022/9/13
Y1 - 2022/9/13
N2 - Von Willebrand factor (VWF) is a multimeric plasma glycoprotein that is critically involved in hemostasis. Biosynthesis of long VWF concatemers in the endoplasmic reticulum and the trans-Golgi is still not fully understood. We use the single-molecule force spectroscopy technique magnetic tweezers to analyze a previously hypothesized conformational change in the D9D3 domain crucial for VWF multimerization. We find that the interface formed by submodules C8-3, TIL3, and E3 wrapping around VWD3 can open and expose 2 buried cysteines, Cys1099 and Cys1142, that are vital for multimerization. By characterizing the conformational change at varying levels of force, we can quantify the kinetics of the transition and stability of the interface. We find a pronounced destabilization of the interface on lowering the pH from 7.4 to 6.2 and 5.5. This is consistent with initiation of the conformational change that enables VWF multimerization at the D9D3 domain by a decrease in pH in the trans-Golgi network and Weibel-Palade bodies. Furthermore, we find a stabilization of the interface in the presence of coagulation factor VIII, providing evidence for a previously hypothesized binding site in submodule C8-3. Our findings highlight the critical role of the D9D3 domain in VWF biosynthesis and function, and we anticipate our methodology to be applicable to study other, similar conformational changes in VWF and beyond.
AB - Von Willebrand factor (VWF) is a multimeric plasma glycoprotein that is critically involved in hemostasis. Biosynthesis of long VWF concatemers in the endoplasmic reticulum and the trans-Golgi is still not fully understood. We use the single-molecule force spectroscopy technique magnetic tweezers to analyze a previously hypothesized conformational change in the D9D3 domain crucial for VWF multimerization. We find that the interface formed by submodules C8-3, TIL3, and E3 wrapping around VWD3 can open and expose 2 buried cysteines, Cys1099 and Cys1142, that are vital for multimerization. By characterizing the conformational change at varying levels of force, we can quantify the kinetics of the transition and stability of the interface. We find a pronounced destabilization of the interface on lowering the pH from 7.4 to 6.2 and 5.5. This is consistent with initiation of the conformational change that enables VWF multimerization at the D9D3 domain by a decrease in pH in the trans-Golgi network and Weibel-Palade bodies. Furthermore, we find a stabilization of the interface in the presence of coagulation factor VIII, providing evidence for a previously hypothesized binding site in submodule C8-3. Our findings highlight the critical role of the D9D3 domain in VWF biosynthesis and function, and we anticipate our methodology to be applicable to study other, similar conformational changes in VWF and beyond.
UR - http://www.scopus.com/inward/record.url?scp=85138117879&partnerID=8YFLogxK
U2 - 10.1182/bloodadvances.2022006978
DO - 10.1182/bloodadvances.2022006978
M3 - Article
C2 - 36069828
AN - SCOPUS:85138117879
SN - 2473-9529
VL - 6
SP - 5198
EP - 5209
JO - Blood advances
JF - Blood advances
IS - 17
ER -