Force Sensing by the Vascular Protein Von Willebrand Factor is Tuned by a Strong Intermonomer Interaction

Mueller, Jochen P.; Mielke, Salomé; Löf, Agneta; Obser, Tobias; Beer, Christof Gerhard; Pippig, Diana A.; Vanderlinden, Willem; Schneppenheim, Reinhard; Benoit, Martin

doi:10.1016/j.bpj.2015.11.212

Cited by 26 publications

(78 citation statements)

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“…In most cases, we observed two unfolding and two refolding steps in the recorded high and moderate force traces, respectively, with extension values matching the expected values for unfolding of the A2 domains (≈180 aa each) that were previously probed in isolation in OT (8,51). Observation of domain (un-)folding only for the two A2 domains is consistent with the prediction that all domains of VWF except A2 are protected against unfolding by long-range disulfide bonds (50,52) and with the results of recent AFM studies (9,10). In addition to the steps attributed to A2 unfolding and refolding, we less frequently also observed larger steps ( Supplementary Fig.…”

Section: Force Clamp Measurements On Full-length Vwf Dimerssupporting

confidence: 89%

“…S6; 70-80 nm at ~11 pN), which we attribute to the dissociation of a strong intermonomer interaction mediated by the D4 domains that has recently been identified in AFM force measurements in approximately one-half of all VWF dimers under near-physiologic conditions (9,10). Consistent with their assignment to the D4mediated intermonomer interaction, the large unfolding steps occur much less frequently in the absence of divalent ions, which have been shown to be critical for the intermonomer interaction (9,10), and are absent for mutant constructs lacking the D4 domain (delD4; Supplementary Fig. S6,S7).…”

Section: Force Clamp Measurements On Full-length Vwf Dimerssupporting

confidence: 52%

“…Since hydrodynamic peak forces grow as the square of the contour length (5,8), transitions that release contour length at low forces are expected to be particularly relevant for VWF's physiological function as they will set of a cascade of increasing forces that trigger additional transitions with further contour length release. Recent work using AFM imaging has suggested transitions in the VWF C-domain stem that, however, could not be detected in AFM-based force spectroscopy, due to its limited force resolution (9,10). Most insights into the mechanical properties and regulation of proteins and their complexes at the single-molecule level have been obtained from force spectroscopy experiments using atomic force microscopes (AFM) or optical tweezers (OT).…”

Section: Introductionmentioning

confidence: 99%

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Modular, ultra-stable, and highly parallel protein force spectroscopy in magnetic tweezers using peptide linkers

Löf

Sedlak

et al. 2018

Preprint

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Single-molecule force spectroscopy has provided unprecedented insights into protein folding, force-regulation, and function. Here, we present a modular magnetic tweezers force spectroscopy approach that uses elastin-like polypeptide linkers to provide a high yield of protein tethers. Our approach extends protein force spectroscopy into the low force (<1 pN) regime and enables ultra-stable measurements on many molecules in parallel. We present (un-)folding data for the single protein domain ddFLN4 and for the large multi-domain dimeric protein von Willebrand factor (VWF) that is critically involved in primary hemostasis. The measurements reveal exponential force-dependencies of unfolding and refolding rates, directly resolve the stabilization of the VWF A2 domain by Ca 2+ , and discover transitions in the VWF C-domain stem at low forces that likely constitute the first steps of VWF activation in vivo. Our modular attachment approach will enable precise and multiplexed force spectroscopy measurements for a wide range of proteins in the physiologically relevant force regime.

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Section: Force Clamp Measurements On Full-length Vwf Dimerssupporting

confidence: 89%

Section: Force Clamp Measurements On Full-length Vwf Dimerssupporting

confidence: 52%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Modular, ultra-stable, and highly parallel protein force spectroscopy in magnetic tweezers using peptide linkers

Löf

Sedlak

et al. 2018

Preprint

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“…It should however, be stressed that to date no study has been published that showed direct interaction of A1 and A2 within a multimer, nor has such an interaction been comprehensively characterized experimentally at the single‐molecule level. When investigating intramolecular interactions within VWF dimers by performing single‐molecule AFM force measurements, Müller, Löf, et al (); Müller, Mielke, et al () did not detect any strong interaction between these domains. Also in optical tweezers experiments by Ying, Ling, Westfield, Sadler, and Shao () on a construct containing three repeats of domains A1, A2, and A3, no such interaction could be found.…”

Section: Resultsmentioning

confidence: 99%

“…It has been shown that VWF multimers adopt a loosely collapsed, globular conformation in solution (Schneider et al, ). The precise nature of the interactions that promote this conformation is not yet well understood, but a recently identified strong intra‐dimer interaction mediated by VWF's D4 domain can be assumed to contribute by promoting a compact conformation of the individual dimers within a multimer (Müller, Löf, et al, , Müller, Mielke, et al, ). Very likely, also additional interactions between the different dimers of a multimer are involved.…”

Section: Vwf A1 Domain Shielding By Intramolecular Interactionsmentioning

confidence: 99%

A biophysical view on von Willebrand factor activation

Löf

Müller

Brehm

2017

Journal Cellular Physiology

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The process of hemostatic plug formation at sites of vascular injury crucially relies on the large multimeric plasma glycoprotein von Willebrand factor (VWF) and its ability to recruit platelets to the damaged vessel wall via interaction of its A1 domain with platelet GPIbα. Under normal blood flow conditions, VWF multimers exhibit a very low binding affinity for platelets. Only when subjected to increased hydrodynamic forces, which primarily occur in connection with vascular injury, VWF can efficiently bind to platelets. This force-regulation of VWF's hemostatic activity is not only highly intriguing from a biophysical perspective, but also of eminent physiological importance. On the one hand, it prevents undesired activity of VWF in intact vessels that could lead to thromboembolic complications and on the other hand, it enables efficient VWF-mediated platelet aggregation exactly where needed. Here, we review recent studies that mainly employed biophysical approaches in order to elucidate the molecular mechanisms underlying the complex mechano-regulation of the VWF-GPIbα interaction. Their results led to two main hypotheses: first, intramolecular shielding of the A1 domain is lifted upon force-induced elongation of VWF; second, force-induced conformational changes of A1 convert it from a low-affinity to a high-affinity state. We critically discuss these hypotheses and aim at bridging the gap between the large-scale behavior of VWF as a linear polymer in hydrodynamic flow and the detailed properties of the A1-GPIbα bond at the single-molecule level.

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Structural hierarchy of mechanical extensibility in human von Willebrand factor multimers

et al. 2022

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The von Willebrand factor (VWF) is a multimeric glycoprotein composed of 80-to 120-nm-long protomeric units and plays a fundamental role in mediating platelet function at high shear. The exact nature of the shear-induced structural transitions have remained elusive; uncovering them requires the high-resolution quantitative analysis of gradually extended VWF. Here, we stretched human blood-plasma-derived VWF with molecular combing and analyzed the axial structure of the elongated multimers with atomic force microscopy. Protomers extended through structural intermediates that could be grouped into seven distinct topographical classes. Protomer extension thus progresses through the uncoiling of the C 1-6 domain segment, rearrangements among the N-terminal VWF domains, and unfolding and elastic extension of the A 2 domain. The least and most extended protomer conformations were localized at the ends and the middle of the multimer, respectively, revealing an apparent necking phenomenon characteristic of plastic-material behavior. The structural hierarchy uncovered here is likely to provide a spatial control mechanism to the complex functions of VWF.

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Force Sensing by the Vascular Protein Von Willebrand Factor is Tuned by a Strong Intermonomer Interaction

Cited by 26 publications

References 0 publications

Modular, ultra-stable, and highly parallel protein force spectroscopy in magnetic tweezers using peptide linkers

Modular, ultra-stable, and highly parallel protein force spectroscopy in magnetic tweezers using peptide linkers

A biophysical view on von Willebrand factor activation

Structural hierarchy of mechanical extensibility in human von Willebrand factor multimers

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