Nav: Home

A better understanding of the von Willebrand Factor's A2 domain

May 20, 2019

Under normal, healthy circulatory conditions, the von Willebrand Factor (vWF) keeps to itself. The large and mysterious multimeric glycoprotein moves through the blood, balled up tightly, its reaction sites unexposed. But when significant bleeding occurs, it springs into action, initiating the clotting process.

When it works properly, vWF helps stop bleeding and saves lives. However, about one to two percent of the world's population is affected by vWF mutations that result in bleeding disorders. For those with more rare, severe forms, a very expensive treatment in the form of blood plasma replacement may be required.

On the other hand, if vWF activates where it isn't needed, it can trigger a stroke or heart attack.

A better understanding of how vWF functions could result in drugs that replace it in those who lack it. It could also lead to the development of new drugs or drug carriers that mimic the protein's behavior for more effective drug delivery.

With that in mind, a team of Lehigh University researchers is working to characterize this mysterious protein. In a recent paper published in Biophysical Journal, they advance experimental data for the shear-induced extensional response of vWF, using a microfluidic device and fluorescence microscopy. Further, they use the results from tandem Brownian dynamics simulations of an experimentally parameterized coarse-grained VWF model to help explain some of their central observations from experiment. This work elucidates further details of the flow-induced biomechanical response behaviors of tethered VWF and demonstrates the power and capabilities of increasingly complex coarse-grained models employed in tandem with experiment.

The paper, called "Shear-Induced Extensional Response Behaviors of Tethered von Willebrand Factor," is authored by Xuanhong Cheng, associate professor of materials science and engineering; Alparslan Oztekin, professor of mechanical engineering and mechanics; Edmund Webb III , associate professor of mechanical engineering and mechanics; and Frank Zhang, associate professor of bioengineering and mechanical engineering and mechanics; as well as doctoral students Michael Morabito and Yi Wang.

vWF at Work

At the location of a minor wound, platelets adhere to the collagen-exposed sites near the hole in the blood vessel wall on their own and act as a plug, effectively stopping the bleeding. Rapid blood flow, however, makes it difficult for platelets to do this. Fortunately, the von Willebrand Factor recognizes this rapid blood flow and activates: "It's a flow-mechanics-activated event, if you will," explains Webb.

The globular molecule unfolds like a Slinky, stretching to 10 times its original size and exposing its reaction sites. It clings to the broken blood vessel wall, where exposed collagen--the structural protein of the blood vessel wall--attracts platelets. vWF then captures platelets from blood as they flow by, acting like a bridge between the collagen and the platelets.

Although the biological function of vWF has long been recognized by scientists, not much is known about the specifics of how vWF functions, particularly under flow conditions.

"Most proteins in blood functions are executed through biochemical reactions," says Cheng. "This protein [vWF] also requires some biochemical reaction for its function, so it needs to grab onto platelets, grab onto collagen--those are biochemical reactions. At the same time, vWF relies on mechanical stimulation to execute the biochemical function, and that part is not very well known. That's what we're trying to study."

Adds Webb: "Some of the data that's coming out of our group but also from other groups indicates that those biochemical reactions are somehow abetted by there being some sort of a tension, a pulling force. So even the biochemical reactions appear to be somewhat mechanically mediated. Again, it was understood that there was this change from a compact, almost ball-like shape, if you will, to this long, stringy thing. But very recently people have been indicating it's not just that. For this chemical site to be active, you have to be pulling it, you have to be in a bit of tension, locally. So it's a really fascinating system."

Unraveling A2

The von Willebrand Factor is a particularly large protein made up of many monomers, or molecules that can be bonded to other identical molecules to form a polymer. Within each monomer of vWF are different domains: A, C and D. Each domain and each of its respective subdomains has its own role, and many of these roles are yet unknown. The A1 domain, for example, binds vWF to platelets. A3 binds vWF to collagen. The A2 domain unfolds to expose the protein's reaction sites, and, when fully opened, exposes a site that permits scission of the vWF molecule down to size. Members of the team have focused on the A2 domain, in particular.

"Understanding that domain and how it interacts with the flow, I think, is the best contribution from our group," says Oztekin.

Each member of the team plays a particular role. Cheng, Zhang and their graduate students work on the experimental side of the project; Oztekin, Webb and their graduate students focus on simulation. Each team's results inform the work of the other.

Zhang, who has been studying vWF for years and brought the project to Lehigh, specializes in single-molecule force spectroscopy and mechanosensing, or how cells respond to mechanical stimuli. He uses a specialized tool called optical tweezers, which utilizes a focused laser beam to apply force to objects as small as a single atom.

"Optical tweezers can grab tiny objects," Zhang explains. "We can grab the vWF and at the same time we apply force to see how the protein changes shape, to see how the proteins are activated when there's a mechanical perturbation or a mechanical force."

Cheng develops microfluidic devices, which have a small diameter and can be used to analyze live bioparticles. She and her team make very small channels similar to the geometry of blood vessels--on the order of 10 micron in height, a few millimeters in length and width--so they can mimic the flow condition that vWF encounters in the body. They tag the vWF molecule fluorescently and use a confocal microscope to capture video and still images of the molecule as it flows through the channel at different rates.

"When we talk about this protein under normal flow, it's one conformation, and then when it's exposed to certain abnormal flow patterns, you'll have a different conformation," Cheng explains. "So we're trying to characterize or replicate that process in an in vitro system, trying to observe how this protein changes conformation under different flow patterns. And then, if we have mutants versus normal protein, how would they behave differently?"

Doctoral student Yi Wang works with Cheng on the microfluidics channel in which they can observe the vWF molecule unraveling and folding back again in real time under a microscope. To do so, they must create an environment that mimics the shear rate, or change in blood flow velocity, found in the body.

"Because we are using a pretty high shear rate to be comparable to the physiological environment, and because of the limited moving speed of a microscope lens that images the molecule, it's actually pretty challenging to capture the movement of a molecule if it's moving," says Wang.

To solve that problem, the team binds one side of the molecule to the surface of the channel to immobilize it as they apply shear force. They have successfully captured the unfolding phenomenon on video.

"If it [the molecule] is bound too tight, it will just stay there [and not unfold]," says Wang. "If it is too loose, everything will be flushed away. So I was very excited when we got the sweet spot of binding it right there on the surface and so it can unfold and fold back."
-end-
This work was supported in part by National Science Foundation grant DMS-1463234 and utilized the Extreme Science and Engineering Discovery Environment, which is supported by National Science Foundation Grant No. ACI-1548562.

ad more about this unique collaboration here: https://www2.lehigh.edu/news/unraveling-vwf-a-better-understanding-of-the-von-willebrand-factors-a2-domain.

Lehigh University

Related Engineering Articles:

Engineering the meniscus
Damage to the meniscus is common, but there remains an unmet need for improved restorative therapies that can overcome poor healing in the avascular regions.
Artificially engineering the intestine
Short bowel syndrome is a debilitating condition with few treatment options, and these treatments have limited efficacy.
Reverse engineering the fireworks of life
An interdisciplinary team of Princeton researchers has successfully reverse engineered the components and sequence of events that lead to microtubule branching.
New method for engineering metabolic pathways
Two approaches provide a faster way to create enzymes and analyze their reactions, leading to the design of more complex molecules.
Engineering for high-speed devices
A research team from the University of Delaware has developed cutting-edge technology for photonics devices that could enable faster communications between phones and computers.
Breakthrough in blood vessel engineering
Growing functional blood vessel networks is no easy task. Previously, other groups have made networks that span millimeters in size.
Next-gen batteries possible with new engineering approach
Dramatically longer-lasting, faster-charging and safer lithium metal batteries may be possible, according to Penn State research, recently published in Nature Energy.
What can snakes teach us about engineering friction?
If you want to know how to make a sneaker with better traction, just ask a snake.
Engineering a plastic-eating enzyme
Scientists have engineered an enzyme which can digest some of our most commonly polluting plastics, providing a potential solution to one of the world's biggest environmental problems.
A new way to do metabolic engineering
University of Illinois researchers have created a novel metabolic engineering method that combines transcriptional activation, transcriptional interference, and gene deletion, and executes them simultaneously, making the process faster and easier.
More Engineering News and Engineering Current Events

Trending Science News

Current Coronavirus (COVID-19) News

Top Science Podcasts

We have hand picked the top science podcasts of 2020.
Now Playing: TED Radio Hour

Teaching For Better Humans 2.0
More than test scores or good grades–what do kids need for the future? This hour, TED speakers explore how to help children grow into better humans, both during and after this time of crisis. Guests include educators Richard Culatta and Liz Kleinrock, psychologist Thomas Curran, and writer Jacqueline Woodson.
Now Playing: Science for the People

#556 The Power of Friendship
It's 2020 and times are tough. Maybe some of us are learning about social distancing the hard way. Maybe we just are all a little anxious. No matter what, we could probably use a friend. But what is a friend, exactly? And why do we need them so much? This week host Bethany Brookshire speaks with Lydia Denworth, author of the new book "Friendship: The Evolution, Biology, and Extraordinary Power of Life's Fundamental Bond". This episode is hosted by Bethany Brookshire, science writer from Science News.
Now Playing: Radiolab

Dispatch 3: Shared Immunity
More than a million people have caught Covid-19, and tens of thousands have died. But thousands more have survived and recovered. A week or so ago (aka, what feels like ten years in corona time) producer Molly Webster learned that many of those survivors possess a kind of superpower: antibodies trained to fight the virus. Not only that, they might be able to pass this power on to the people who are sick with corona, and still in the fight. Today we have the story of an experimental treatment that's popping up all over the country: convalescent plasma transfusion, a century-old procedure that some say may become one of our best weapons against this devastating, new disease.   If you have recovered from Covid-19 and want to donate plasma, national and local donation registries are gearing up to collect blood.  To sign up with the American Red Cross, a national organization that works in local communities, head here.  To find out more about the The National COVID-19 Convalescent Plasma Project, which we spoke about in our episode, including information on clinical trials or plasma donation projects in your community, go here.  And if you are in the greater New York City area, and want to donate convalescent plasma, head over to the New York Blood Center to sign up. Or, register with specific NYC hospitals here.   If you are sick with Covid-19, and are interested in participating in a clinical trial, or are looking for a plasma donor match, check in with your local hospital, university, or blood center for more; you can also find more information on trials at The National COVID-19 Convalescent Plasma Project. And lastly, Tatiana Prowell's tweet that tipped us off is here. This episode was reported by Molly Webster and produced by Pat Walters. Special thanks to Drs. Evan Bloch and Tim Byun, as well as the Albert Einstein College of Medicine.  Support Radiolab today at Radiolab.org/donate.