The N-terminal autoinhibitory module of the A1 domain in von Willebrand factor stabilizes the mechanosensor catch bond

The von Willebrand factor (VWF), by interacting with the circulatory system and platelets, harnesses hemodynamic forces to form hemostatic plugs or occlusive thrombi. The autoinhibitory modules (AIMs) flanking the VWF-A1 domain were found to contribute to its biomechanical activation. However, how AIM sequences regulate the VWF-A1 binding behavior is controversial and incompletely understood as their structures are currently unsolvable by crystallography. To address this, we first performed molecular dynamics simulations to predict the N-terminal AIM (N-AIM; residues Q1238-E1260) structure. Excitingly, we found that N-AIM could cooperate with C-AIM to form a joint Rotini-like structure, thereby partially autoinhibiting the VWF-A1-GPIb alpha interaction. Furthermore, we used biomembrane force probe (BFP) assays to experimentally demonstrate that the VWF-A1 containing long N-AIM sequence (1238-A1) exhibited catch-bond behavior as the force first decelerated (catch) and then accelerated (slip) the dissociation. Conversely, VWF-A1 with short N-AIM (1261-A1) displayed bi-variable behaviors with either catch (1261(H)-A1) or slip bonds (1261(L)-A1). Notably, such bi-variable transition happened at low temperatures or high pH levels, whereas Q1238-E1260 stabilized the 1238-A1 catch bond regardless of the environmental factors. The physiological study was complemented by platelet perfusion assays using microfluidics. Taken together, these studies provide new mechanobiology on how N-AIM serves as a mechano-regulator of VWF activity, which inspires future VWF-A1 dependent antithrombotic approaches.

Automated summary

The von Willebrand factor (VWF) forms hemostatic plugs or occlusive thrombi by interacting with the circulatory system and platelets. This study focuses on the role of autoinhibitory modules (AIMs) in regulating VWF-A1 binding behavior. Molecular dynamics simulations and biomembrane force probe (BFP) assays reveal the structure and catch-bond behavior of the N-terminal AIM (N-AIM), providing insight into how it serves as a mechano-regulator of VWF activity. Platelet perfusion assays using microfluidics further support these findings. Overall, this research sheds light on new mechanobiology of VWF activity and inspires antithrombotic approaches targeting VWF-A1.