Elongational flow induces the unfolding of von Willebrand factor at physiological flow rates.
Ontology highlight
ABSTRACT:
SUBMITTER: Sing CE
PROVIDER: S-EPMC2862189 | biostudies-literature | 2010 May
REPOSITORIES: biostudies-literature
Publications
Elongational flow induces the unfolding of von Willebrand factor at physiological flow rates.
Sing Charles E CE Alexander-Katz Alfredo A
Biophysical journal 20100501 9
The unfolding of von Willebrand Factor (vWF), one of the largest multimeric proteins in our body, has been shown to be a crucial step in the process of blood clotting. Here we show that elongational flows, which appear during vasoconstriction or stenosis, are the primary activation mechanisms of vWF, and unfold the multimeric protein at flow rates that are two orders-of -magnitude below those corresponding to pure shear. The findings presented here complement the current understanding of blood c ...[more]
Similar Datasets
Project description:The globular-to-unraveled conformation transition of von Willebrand factor (vWF), a large polymeric glycoprotein in human blood plasma, is a crucial step in the process of clotting at sites of vascular injury. However, unraveling of vWF multimers in uninjured vasculature can lead to pathology (i.e., thrombus formation or degradation of vWF proteins by enzyme ADAMTS13, making them nonfunctional). To identify blood flow conditions that might induce pathological unraveling of vWF multimers, here we have computed the globular-to-unraveled transition rate of vWF multimers subjected to varying strain rate elongational flow by employing an enhanced sampling technique, the weighted ensemble method. Weighted ensemble sampling was employed instead of standard brute-force simulations because pathological blood flow conditions can induce undesired vWF unraveling on timescales potentially inaccessible to standard simulation methods. Results here indicate that brief but periodic exposure of vWF to the elongational flow of strain rate greater than or equal to 2500 s-1 represents a source of possible pathology caused by the undesired unraveling of vWF multimers.
| S-EPMC8204344 | biostudies-literature
Project description:Using Brownian molecular dynamics simulations, we examine the internal dynamics and biomechanical response of von Willebrand factor (vWF) multimers subject to shear flow. The coarse grain multimer description employed here is based on a monomer model in which the A2 domain of vWF is explicitly represented by a nonlinear elastic spring whose mechanical response was fit to experimental force/extension data from vWF monomers. This permits examination of the dynamic behavior of hydrodynamic forces acting on A2 domains as a function of shear rate and multimer length, as well as position of an A2 domain along the multimer contour. Force/position data reveal that collapsed multimers exhibit a force distribution with two peaks, one near each end of the chain; unraveled multimers, however, show a single peak in A2 domain force near the center of multimers. Guided further by experimental data, significant excursions of force acting on a domain are associated with an increasing probability for A2 domain unfolding. Our results suggest that the threshold shear rate required to induce A2 domain unfolding is inversely proportional to multimer length. By examining data for the duration and location of significant force excursions, convincing evidence is advanced that unfolding of A2 domains, and therefore scission of vWF multimers by the size-regulating blood enzyme ADAMTS13, happen preferentially near the center of unraveled multimers.
| S-EPMC6303231 | biostudies-literature
Project description:von Willebrand Factor is a mechano-sensitive protein circulating in blood that mediates platelet adhesion to subendothelial collagen and platelet aggregation at high shear rates. Its hemostatic function and thrombogenic effect, as well as susceptibility to enzymatic cleavage, are regulated by a conformational change from a collapsed globular state to a stretched state. Therefore, it is essential to account for the conformation of the vWF multimers when modeling vWF-mediated thrombosis or vWF degradation. We introduce a continuum model of vWF unfolding that is developed within the framework of our multi-constituent model of platelet-mediated thrombosis. The model considers two interconvertible vWF species corresponding to the collapsed and stretched conformational states. vWF unfolding takes place via two regimes: tumbling in simple shear and strong unfolding in flows with dominant extensional component. These two regimes were demonstrated in a Couette flow between parallel plates and an extensional flow in a cross-slot geometry. The vWF unfolding model was then verified in several microfluidic systems designed for inducing high-shear vWF-mediated thrombosis and screening for von Willebrand Disease. The model predicted high concentration of stretched vWF in key regions where occlusive thrombosis was observed experimentally. Strong unfolding caused by the extensional flow was limited to the center axis or middle plane of the channels, whereas vWF unfolding near the channel walls relied upon the shear tumbling mechanism. The continuum model of vWF unfolding presented in this work can be employed in numerical simulations of vWF-mediated thrombosis or vWF degradation in complex geometries. However, extending the model to 3-D arbitrary flows and turbulent flows will pose considerable challenges.
| S-EPMC9847011 | biostudies-literature
Project description:Von Willebrand factor (VWF) is a multimeric plasma glycoprotein involved in both hemostasis and thrombosis. VWF conformational changes, especially unfolding of the A2 domain, may be required for efficient enzymatic cleavage in vivo. It has been shown that a single A2 domain unfolds at most probable unfolding forces of 7-14 pN at force loading rates of 0.35-350 pN/s and A2 unfolding facilitates A2 cleavage in vitro. However, it remains unknown how much force is required to unfold the A2 domain in the context of a VWF multimer where A2 may be stabilized by other domains like A1 and A3. With the optical trap, we stretched VWF multimers and a poly-protein (A1A2A3)3 that contains three repeats of the triplet A1A2A3 domains at constant speeds of 2000 nm/s and 400 nm/s, respectively, which yielded corresponding average force loading rates of 90 and 22 pN/s. We found that VWF multimers became stiffer when they were stretched and extended by force. After force increased to a certain level, sudden extensional jumps that signify domain unfolding were often observed. Histograms of the unfolding force and the unfolded contour length showed two or three peaks that were integral multiples of approximately 21 pN and approximately 63 nm, respectively. Stretching of (A1A2A3)3 yielded comparable distributions of unfolding force and unfolded contour length, showing that unfolding of the A2 domain accounts for the behavior of VWF multimers under tension. These results show that the A2 domain can be indeed unfolded in the presence of A1, A3, and other domains. Compared with the value in the literature, the larger most probable unfolding force measured in this study suggests that the A2 domain is mechanically stabilized by A1 or A3 although variations in experimental setups and conditions may complicate this interpretation.
| S-EPMC2856187 | biostudies-literature
Project description:Essentials von Willebrand factor (VWF) function is shear stress dependent. Platelet accumulation in a microfluidic assay correlates with VWF levels. The microfluidic assay discriminates type 1 von Willebrand disease from healthy controls. The microfluidic flow assay detects responses to therapeutic intervention (DDAVP).SummaryBackground von Willebrand disease (VWD) is a mucocutaneous bleeding disorder with a reported prevalence of 1 in 10 000. von Willebrand factor (VWF) function and platelet adhesion are regulated by hemodynamic forces that are not integrated into most current clinical assays. Objective We evaluated whether a custom microfluidic flow assay (MFA) can screen for deficiencies in VWF in patients presenting with mucocutaneous bleeding. Methods Whole blood from individuals with mucocutaneous bleeding was assayed in a custom MFA. Results Thirty-two patients with type 1 VWD (10/32) or reported mucocutaneous bleeding were enrolled. The platelet adhesion velocity (r = 0.5978 for 750 s-1 and 0.6895 for 1500 s-1 ) and the maximum platelet surface area coverage (r = 0.5719 for 750 s-1 and 0.6633 for 1500 s-1 ) in the MFA correlated with VWF levels. Furthermore, the platelet adhesion velocity at 750 s-1 (type 1 VWD, mean 0.0009761, 95% confidence interval [CI] 0.0003404-0.001612; control, mean 0.003587, 95% CI 0.002455-0.004719) and at 1500 s-1 (type 1 VWD, mean 0.0003585, 95% CI 0.00003914-0.0006778; control, mean 0.003132, 95% CI 0.001565-0.004699) differentiated type 1 VWD from controls. Maximum platelet surface area coverage at 750 s-1 (type 1 VWD, mean 0.1831, 95% CI 0.03816-0.3281; control, mean 0.6755, 95% CI 0.471-0.88) and at 1500 s-1 (type 1 VWD, mean 0.07873, 95% CI 0.01689-0.1406; control, mean 0.6432, 95% CI 0.3607-0.9257) also differentiated type 1 VWD from controls. We also observed an improvement in platelet accumulation after 1-desamino-8-d-arginine vasopressin (DDAVP) treatment at 1500 s-1 (pre-DDAVP, mean 0.4784, 95% CI 0.1777-0.7791; post-DDAVP, mean 0.8444, 95% CI 0.7162-0.9726). Conclusions These data suggest that this approach can be used as a screening tool for VWD.
| S-EPMC5794217 | biostudies-literature
Project description:Unusually large von Willebrand factor (VWF), the first responder to vascular injury in primary hemostasis, is designed to capture platelets under the high shear stress of rheological blood flow. In type 2M von Willebrand disease, two rare mutations (G1324A and G1324S) within the platelet GPIb? binding interface of the VWF A1 domain impair the hemostatic function of VWF. We investigate structural and conformational effects of these mutations on the A1 domain's efficacy to bind collagen and adhere platelets under shear flow. These mutations enhance the thermodynamic stability, reduce the rate of unfolding, and enhance the A1 domain's resistance to limited proteolysis. Collagen binding affinity is not significantly affected indicating that the primary stabilizing effect of these mutations is to diminish the platelet binding efficiency under shear flow. The enhanced stability stems from the steric consequences of adding a side chain (G1324A) and additionally a hydrogen bond (G1324S) to His(1322) across the ?2-?3 hairpin in the GPIb? binding interface, which restrains the conformational degrees of freedom and the overall flexibility of the native state. These studies reveal a novel rheological strategy in which the incorporation of a single glycine within the GPIb? binding interface of normal VWF enhances the probability of local unfolding that enables the A1 domain to conformationally adapt to shear flow while maintaining its overall native structure.
| S-EPMC4759165 | biostudies-literature
Project description:Von Willebrand factor (VWF) activates in response to shear flow to initiate hemostasis, while aberrant activation could lead to thrombosis. Above a critical shear force, the A1 domain of VWF becomes activated and captures platelets via the GPIb-IX complex. Here we show that the shear-responsive element controlling VWF activation resides in the discontinuous autoinhibitory module (AIM) flanking A1. Application of tensile force in a single-molecule setting induces cooperative unfolding of the AIM to expose A1. The AIM-unfolding force is lowered by truncating either N- or C-terminal AIM region, type 2B VWD mutations, or binding of a ristocetin-mimicking monoclonal antibody, all of which could activate A1. Furthermore, the AIM is mechanically stabilized by the nanobody that comprises caplacizumab, the only FDA-approved anti-thrombotic drug to-date that targets VWF. Thus, the AIM is a mechano-regulator of VWF activity. Its conformational dynamics may define the extent of VWF autoinhibition and subsequent activation under force.
| S-EPMC8060278 | biostudies-literature
Project description:von Willebrand Factor (vWF) is a large multimeric protein that binds to platelets and collagen in blood clotting. vWF A2 domain hosts a proteolytic site for ADAMTS13 (A Disintegrin and Metalloprotease with a ThromboSpondin type 1 motif, member 13) to regulate the size of vWF multimers. This regulation process is highly sensitive to force conditions and protein-glycan interactions as the process occurs in flowing blood. There are two sites on A2 domain (N1515 and N1574) bearing various N-linked glycan structures. In this study, we used molecular dynamics (MD) simulation to study the force-induced unfolding of A2 domain with and without a single N-linked glycan type on each site. The sequential pullout of β-strands was used to represent a characteristic unfolding sequence of A2. This unfolding sequence varied due to protein-glycan interactions. The force-extension and total energy-extension profiles also show differences in magnitude but similar characteristic shapes between the systems with and without glycans. Systems with N-linked glycans encountered higher energy barriers for full unfolding and even for unfolding up to the point of ADAMTS13 cleavage site exposure. Interestingly, there is not much difference observed for A2 domain structure itself with and without glycans from standard MD simulations, suggesting roles of N-glycans in A2 unfolding through long-ranged protein-glycan interactions.
| S-EPMC6207679 | biostudies-literature
Project description:Streptococcus pneumoniae is a major cause of community acquired pneumonia and septicaemia in humans. These diseases are frequently associated with thromboembolic cardiovascular complications. Pneumococci induce the exocytosis of endothelial Weibel-Palade Bodies and thereby actively stimulate the release of von Willebrand factor (VWF), which is an essential glycoprotein of the vascular hemostasis. Both, the pneumococcus induced pulmonary inflammation and the thromboembolytic complications are characterized by a dysbalanced hemostasis including a marked increase in VWF plasma concentrations. Here, we describe for the first time VWF as a novel interaction partner of capsulated and non-encapsulated pneumococci. Moreover, cell culture infection analyses with primary endothelial cells characterized VWF as bridging molecule that mediates bacterial adherence to endothelial cells in a heparin-sensitive manner. Due to the mechanoresponsive changes of the VWF protein conformation and multimerization status, which occur in the blood stream, we used a microfluidic pump system to generate shear flow-induced multimeric VWF strings on endothelial cell surfaces and analyzed attachment of RFP-expressing pneumococci in flow. By applying immunofluorescence visualization and additional electron microscopy, we detected a frequent and enduring bacterial attachment to the VWF strings. Bacterial attachment to the endothelium was confirmed in vivo using a zebrafish infection model, which is described in many reports and acknowledged as suitable model to study hemostasis mechanisms and protein interactions of coagulation factors. Notably, we visualized the recruitment of zebrafish-derived VWF to the surface of pneumococci circulating in the blood stream and detected a VWF-dependent formation of bacterial aggregates within the vasculature of infected zebrafish larvae. Furthermore, we identified the surface-exposed bacterial enolase as pneumococcal VWF binding protein, which interacts with the VWF domain A1 and determined the binding kinetics by surface plasmon resonance. Subsequent epitope mapping using an enolase peptide array indicates that the peptide 181YGAEIFHALKKILKS195 might serve as a possible core sequence of the VWF interaction site. In conclusion, we describe a VWF-mediated mechanism for pneumococcal anchoring within the bloodstream via surface-displayed enolase, which promotes intravascular bacterial aggregation.
| S-EPMC6443961 | biostudies-literature
Project description:Von Willebrand factor, an ultralarge concatemeric blood protein, must bind to platelet GPIbα during bleeding to mediate hemostasis, but not in the normal circulation to avoid thrombosis. Von Willebrand factor is proposed to be mechanically activated by flow, but the mechanism remains unclear. Using microfluidics with single-molecule imaging, we simultaneously monitored reversible Von Willebrand factor extension and binding to GPIbα under flow. We show that Von Willebrand factor is activated through a two-step conformational transition: first, elongation from compact to linear form, and subsequently, a tension-dependent local transition to a state with high affinity for GPIbα. High-affinity sites develop only in upstream regions of VWF where tension exceeds ~21 pN and depend upon electrostatic interactions. Re-compaction of Von Willebrand factor is accelerated by intramolecular interactions and increases GPIbα dissociation rate. This mechanism enables VWF to be locally activated by hydrodynamic force in hemorrhage and rapidly deactivated downstream, providing a paradigm for hierarchical mechano-regulation of receptor-ligand binding.Von Willebrand factor (VWF) is a blood protein involved in clotting and is proposed to be activated by flow, but the mechanism is unknown. Here the authors show that VWF is first converted from a compact to linear form by flow, and is subsequently activated to bind GPIbα in a tension-dependent manner.
| S-EPMC5567343 | biostudies-literature