HDX for understanding a protein’s higher order structure and function | Behind the Science

HDX for understanding a protein’s higher order structure and function | Behind the Science


Welcome to another episode
of Behind the Science– On Location. I’m your host,
Jennifer Fournier. One of many challenges that
protein scientists have been trying to
tackle for centuries now is understanding
just how proteins interact in the complex
human body universe. Let’s go to one of
the original labs here at Waters that helped
to develop a cool solution to this hot topic Hi, Keith. You are just the person
I was looking for. Hey, Jen, how’s it going? Good. So I am working on
proteins, and I’m trying to understand
protein-protein interaction in the human body,
and I was wondering if there are any techniques
that I could use? Well, one of the
biggest ones out there– and it’s been out
for a long time– is X-ray crystallography. It’s a great technique. You can really deduce
a lot of things about protein
structure that way. Even some binding experiments
can be done that way. But it really has
one major drawback. So what is that drawback? Well, really, it’s a
snapshot in time and space. Really, if we’re looking at
how that protein interacts in your body–
physiological conditions, you really can’t tell that
with X-ray crystallography. Yeah, I really need to
understand the interaction within the body. So are there techniques
that I could use for that? I got something for you. It’s actually right here. It’s called hydrogen-deuterium
exchange, or HDX. So I have heard of
HDX, but I’ve also heard that it’s not necessarily
robust or reproducible. That’s a fair point. And it probably was true
about 10, 15 years ago. It was a technique that really
pioneered in academics labs and really was not easily
adoptable in industry– in biopharm. And the biggest reason for
that is because everything was performed in an ice bucket. And if you want
reproducible results, working through an ice bucket
every day in and day out is really not what
you want to be doing. Yeah, I would think that’d
be really difficult. So how are we changing that? Well, again, through
a collaboration we had with folks around the US,
we’ve developed an HDX manager. And this basically is a
glorified refrigerator. It’s a box that has our
valves, our columns, everything inside there– keep everything nice
and cool and stable for the entire experiment
you want to run. So basically, this module
is a glorified ice bucket. Absolutely. Can you take me through
a little bit more how this technique works? I can do one step better, Jen. I actually have a
video I can show you. Sounds great. All right. So Jen, what we have here
is two different flavors of the HDX system. On the left, you
see the mail system, which works fine in
most academics labs. They don’t need the
automation necessary. And on the right is what,
really, our biopharm customers tend to like, and it’s
a full-blown automation with robotics. That’s what you saw
me standing in front of when you came to find me. That robotics platform handles
everything– from sample prep, through the workflow, through
the injection to the system, and triggers a mass spec and
all the acquisition steps necessary. So really is a turnkey solution. And really, beyond
that platform– beyond the notion
of the robotics– we’ve also done steps to make every bit of the workflow possible. Again, automation you
see, but also things like digestion, with an enzymate
pepsin digestion column. We also have the LC system,
the ACQUITY UPLC M-Class with the mass spec you saw. But the biggest piece that
we added later in the game was the informatics package– DynamX software. It was great, we found early
on in the collaboration. We did a great thing by
getting people their data much faster, much
more reproducible, but they were stuck
in bottlenecks. They had been working up those
results they wanted to see. And with the advent of DynamX– that informatics package–
we really pioneered that level of workflow. Also in the area are things
like analytical standards– just to test the robustness
and reproducibility of the platform. We want to assure that
every result we give you from start to finish is
consistent and constant with what you’re hoping to get. So Keith, it really sounds like
we’ve revolutionized the way that people are
using HDX technology, and it sounds like our academic
collaborators played a big role in this. That’s absolutely true, Jen. It wouldn’t have been
possible to achieve what we’ve achieved
without that benefit of those collaborations. It was through a series of
both early beta evaluations, through iterative designs
with collaborators, from not only the hardware
perspective, but even the software we
talked about earlier. So what other applications
are people doing with HDX? So biopharm companies
do a lot of things, like epitope mapping. They want to study
where proteins and small molecules that
are approaching the proteins interact. Things like
formulation stability– they want to share that
protein stays in the right form and function during drug
release and drug development. But also, we get
a lot of academics doing very interesting things. Not only is it looking at
Alzheimer’s and the HIV virus, but also a collaborator
in Singapore is looking at a Dengue
virus and seeing what HDX can tell them
about the structure of such an elusive molecule. That is so neat–
though I have to admit that I’m quite relieved,
because when you pointed over to this ice bucket, I really
thought that we were in the ice bucket challenge video,
and I’m so glad that I didn’t get wet today. Well, Jen, it’s still early. You can get some water
in there for sure. No, I think we’re good. Thank you, Keith. You’re welcome. The HDX system is
a full package tool that will get you past one
of the challenges you face– understanding the complexity
of higher-order structure in the human body. If you want to learn more
about this cool technique and see how this tool
advances science, watch the videos at the link
below, and join us next time for another episode of
Behind the Science– On Location.