
Cryo-electron microscopy breaks a key barrier - pseudolus
https://www.nature.com/articles/d41586-020-01658-1
======
fabian2k
The headline is potentially confusing, it doesn't mean that this is the first
time individual atoms were measured, but the first time this particular method
has been pushed to such a high resolution that individual atoms could be
distinguished.

Cryo EM is a very hot method right now to determine the three-dimensional
structure of large molecules like e.g. proteins or protein complexes.
Something like 10-15 years ago the best you could get was maybe 1.5 - 2 nm
(these are rough numbers from memory), the results described in the article
are 0.17 nm.

~~~
jjoonathan
Further, atomic resolution is routine in electron microscopy, it's
specifically Cryo EM (a sub-technique aimed at imaging proteins, which are
extremely fragile compared to, say, chunks of metal or computer chips) where
the advance has happened.

This is phenomenal! The title still feels a bit sensationalized, but don't
they all?

~~~
m4rtink
Cryo EM is really cool (not just in temperature! ;-) ) technology! :) Recently
I had a chance to go on a tour in the two-part CEITEC research center in Brno,
a city with long history of electron microscopy manufacturing and research.

One part of the CEITEC research center does "dry" tech - chemistry, robotics,
motor and machine control and chips. Of course, they have quite a lot of
cutting edge electron microscopes to image various nano structures. All
looking super high tech as you watch their shiny metal over the clean room
window. All the machines stand on concrete slabs embedded into the hill below
the building to avoid vibrations and have ultra high vacuum inside.

The other part of CEITEC does "wet" research - proteins, cells, bacteria,
microorganisms. Of course also they have cutting edge, cry electron
microscopes. These looked quite different to the other ones - it was a huge
black cube, reaching almost the high ceiling of the room. You could not see
all the glistening internal components like in the previous case. In this case
they even let us to the room itself - apparently given all the volatiles in
the samples, the cleanliness requirements in the room containing the machine
are not as strict & it likely (my guess) needs to keep quite a bit part of the
machine at very low temperatures, explaining all the covers. IIRC the software
controlling the camera runs on Linux & engineers from Brno working for one of
the big EM manufacturers were involved in building this enormous cryo EM
machine, which was a pretty nice touch. :)

~~~
hoseja
Small world :)

~~~
Koshkin
Until you start counting them in moles.

------
COGlory
I'm a structural biologist that uses CryoEM to study archeal viruses. I'd be
happy to answer any questions about CryoEM.

I did want to point out though that this title is misleading. The researchers
didn't get an image of an atom. Instead, they reconstructed hundreds of
thousands of images of the protein to determine where the atoms are almost
exactly. So there's no actual image that shows an atom, as cool as that would
be.

~~~
mynegation
What are the limitations of this technique in determining spatial structure of
the proteins? Do they get denaturated when cooled?

~~~
gewa
No, the proteins are vitrified very rapidly in liquid ethane at approx.
-200°C, so that the structure ist conserved.

~~~
tynpeddler
Protein "structure" is a slippery concept. In situ measurements, molecular
dynamics simulations and basic common sense pretty conclusively show that most
proteins actually have a number of conformations they access during normal
biological processes. In many cases, these conformations are required for the
proteins normal functions. I used to work in the de novo protein field and it
was a constant source of irritation to me that what we mainly designed were
crystal structures, not dynamic functional proteins.

~~~
mynegation
Thank you. Would it be fair to say that crystallized protein is always in one
specific confirmation? Compared to that, would vitrification have the chance
(however random) to capture protein molecules in different conformations?

~~~
COGlory
Yes, a crystallized protein will always be in a single conformation, or you
won't be able to see it because the electron density map will be an average of
all possible conformations, and therefore meaningless.

Single particle gives you the opportunity to see different conformations, but
only if the data is discrete. If there's a continuous amount of conformations
(think a molecular motor that's rotating) you would need nearly infinite data
to resolve a nearly infinite number of conformations. If the data is less than
continuous, you can image enough particles to see all the different
conformations by constructing multiple models in parallel and using 3D angular
searching to bin them by what conformation they are in. This is a
computationally exhausting process, however.

~~~
natechols
I hate to be that guy, but your first statement is technically incorrect: you
can have discrete alternate conformations superimposed in the electron density
(usually 2-3 is the most that can be resolved), and you can have different
conformations of multiple copies of the molecule. (I've personally worked with
both, although the differences in the second case were small.) That's not even
counting ensemble-based approaches to modeling the crystal structure, although
that's arguably just a different way of representing the uncertainty.

All that aside, crystallization certainly biases it towards specific
conformations, which single-particle EM does not.

~~~
COGlory
Didn't realize that. My experience with xray crystallography is limited.
Obviously there's some variance, but I always assumed it would simply be
unresolvable/disordered in that case.

------
jgehrcke
While this is a breakthrough within the field of cryo-electron microscopy it
is important to appreciate that for many questions in structural biology we
also need to understand how protein structure changes over time.

With the presented method the structure sampling time seems to be O(10 s)
which easily is about 10 orders of magnitudes slower than the dynamics we're
interested in seeing.

A direct consequence of this is that for achieving atomic spatial resolution
they _needed_ to use a "rock-solid" protein -- one that has an exceptionally
stiff structure (one that does not wiggle a lot). The presented method is
great and cool, but this is a pretty severe limitation. Most proteins wiggle a
lot :-).

Background: protein structure-function relationship can often be well
understood only when considering the structural dynamics of the protein (key
words: conformational changes, the entropic contribution to free energy).

That is, in the ideal case we would be able to measure molecular structure not
only at high spatial resolution, but also at high temporal resolution.

How can one make such a measurement _much_ faster, by ~10 orders of magnitude?
By irradiating a _lot_ of light. Via X-ray free electron lasers (XFEL).

XFEL-based techniques are expected to revolutionize structural biology (as
always, also still a long way to go):

> XFEL protein crystallography not only determines high resolution structures
> of proteins, but also reveals the time-stamped conformational changes of
> proteins.

\-
[https://www.nature.com/articles/nmeth.3070.pdf?origin=ppub](https://www.nature.com/articles/nmeth.3070.pdf?origin=ppub)

\-
[https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6678726/](https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6678726/)

(PhD in structural biology/bioinformatics, investigated dynamics of proteins
with molecular dynamics simulations)

~~~
Ultimatt
You're glossing over the advantages specific to cryo-EM though, which is it
can give you a good picture of large ensembles of small things at this
accuracy, with a lot less computational/interpretation hardship than X-ray
crystallography or NMR. So for seeing structures of large protein complexes
and how the super structure varies with heteromeric variation this is a really
big deal. If you wanted to see what viral capsids in-situ looked like down to
the atoms this is what you'd use. There are a lot of places this is going to
be useful, there is no one method to rule them all in structural proteomics
simply because nothing can offer all of the spatial and temporal scales one
might need. Plus cryo would only give you the surface hull of these complexes,
you'd still need a different method to fill in all the exact structures
inside. Writing up these methods as if there is some competition for "best" is
an entirely false impression to a lay person reading. The best is all methods
improve and researchers collaborate to provide every possible scoped view.
Even mass spectrometry is doing crazy things for investigating the structure
of disordered proteins. You'd never use that for something you would use cryo-
EM for.

~~~
jgehrcke
Thanks for this reply and the additional level of detail. I didn't mean to
imply that one method is generally better than another. The combination of a
variety of methods applied to the same problem space is certainly the way to
go whenever we really want to unravel molecular mechanisms.

------
benjaminva
I had the privilege to ocasionally work with Holger Stark on similar structure
determination challanges and can confirm that they really pushed the limits
here thanks to very smart statistical methods. The resolution heavily depends
on correctly sorting/classifying the large amount (> 10k for sure) images of
these small particles.

There exists a race in the structural biology community about the next big
method that allows to determine structures of proteins that were hard to
crystalize and it seems that CryoEM is becoming the winner in this race.

As an alternative approach, people are building large X-ray lasers that have
extremly high intensity and short pulse lengths which they plan to shoot at
single particles and resolve individual scattering images. This method can
very likely also achieve atomic resolution of non-crystaline particles, even
without the need to freeze them down.

It will be exciting times for the whole bio-chemical physics community.
Congrats to Holger and the team for another great publication in Nature.

~~~
natechols
Having worked in X-ray crystallography on synchrotrons and XFELs, I can't take
the single-particle XFEL concept seriously any more. Ten years ago it seemed
impressive, but the EM technology has advanced so much faster - and it's a
fraction of the cost. There are some neat applications of nanocrystallography,
but that too seems like a niche application now, not a magic bullet for
accelerating protein structure determination.

------
TomJansen
The real breakthrough here is that with this technique you don't have to
crystallize the proteins anymore, which is the biggest bottleneck in X-Ray
Crystallography.

Relevant; they used Cyro-EM to elucidate the structure of the SARS-CoV-2 Spike
protein [1], which is a big step towards the development of a vaccine. This is
a resolution of 3.5 ångström, but still a very nice feat.

[1]:
[https://science.sciencemag.org/content/367/6483/1260](https://science.sciencemag.org/content/367/6483/1260)

~~~
l33tman
3.5Å cryo-EM structures have existed for some time. The important thing coming
down under 2Å or so, is that it drastically improves the quality of the
molecular dynamics simulations you build of these structures.

~~~
natechols
I'm skeptical of molecular dynamics and docking studies... but the ability to
clearly resolve bound drug molecules, or host-pathogen interactions, is very
valuable indeed.

------
iagovar
So, pardon my ignorance, but can we finally say that atoms are particles with
concrete dimensions? Last time I checked the whole field seemed measuring
everything into probabilistic and statistical terms.

~~~
marcosdumay
You can resolve atoms despite all the uncertainty just like you can resolve a
tree at distance despite it not having a clear surface.

The electron distribution falls with an exponential function on large
distances, what is much cleaner than a tree surface anyway.

~~~
AtHeartEngineer
This is a good analogy

------
mysterypie
Could someone confirm what we’re looking at in that picture please? In that
spider web of blue points joined by blue line segments: Those tiny blue points
are atoms? The blue line segments are covalent bonds? And what’s the
difference between the blue and purple areas?

If those line segments are probability distributions of shared electrons, that
means our high school mental model of a covalent bond (drawing a line between
two atoms) is not too bad.

~~~
natechols
The line segments define surfaces around the regions of highest electron
density. This is how X-ray crystallographers traditionally viewed electron
density, probably because that simple mesh was the best that early computer
graphics systems could handle. In this particular image they've shown nested
surfaces - my guess would be at 1 (purple), 2, and 3 standard deviations above
the mean, or something like that. Again, not atoms, just electron clouds, but
obviously the regions around the atoms have the most electrons.

~~~
mysterypie
So it’s not like what I thought at all. If I understand you correctly, then
each one of those bright blue balls is a cloud of electrons at the center of
which is a nucleus (but we don’t see the nucleus itself). The tiny blue lines
and vertices are just computer generated imagery to define the “surface” of an
atom. Those hexagons with six blue balls joined by purple mesh are benzene
rings? So each purple area defines a cloud of shared electrons in a benzene
ring?

~~~
mncharity
> computer generated imagery to define the “surface” of an atom

The grids are isosurfaces of electron density. Analogous to isolines on a map
illustrating mountain terrain height.

Atoms are sticky little fuzzy balls. The fuzz, electron density, falls off
exponentially. They don't have an "edge" in a non-fuzzy-object sense.
Analogous to a stereotype volcano having an arbitrary perimeter. Where would I
place it? Where you need climbing gear? Where biking up gets hard? In the
foothills? Somewhere on the sloped plain? You place the perimeter wherever is
convenient for what you're trying to use it for. People are usually interested
in the densities between sticking atoms, which are several orders of magnitude
down from peak density. Analogous to drawing an isoline at "the towering
volcano is here high enough to start stubbing your toe on it". The picture
draws isosurfaces of electron density at something like "around here one atom
would 'bump' another, and you can see bonds" and at "I wrap a single atom
only, but still have an interesting shape".

To show the various features of interest, one draws multiple isosurfaces. As
with a terrain map. But an exponential electron density scale is harder to
represent than linear height scale. And it's hard to clearly draw more than a
couple of nested 3D isosurfaces. So instead of placing many iso's at linear
steps, here there are just a couple, more like an order of magnitude apart.

~~~
natechols
The terrain analogy is great - this is how the first structural biologists had
to view electron density: [https://image.slideserve.com/105608/electron-
density-maps-of...](https://image.slideserve.com/105608/electron-density-maps-
of-proteins-sperm-whale-myoglobin-2-4-angstrom-resolution-l.jpg) I think this
is a stack of 2d "slices" through the density, printed on transparencies. But
I've also seen 2d representations of charge density (some of which is just
theoretical studies) that are even more similar to a terrain map.

~~~
mncharity
Neat. I've been exploring visual representations of electron density,
optimized for educational use with XR. Intending to illustrate that science
education content could be made transformatively less wretched.

Isosurfaces and nonlinear color gradients are educationally problematic. And
fuzz gradients are hard to visually stereo fuse. And on near-term AR displays,
black is transparent, and background environment colors can't be manipulated,
which limits the space of attainable colorizations.

My current thought is to approach it as a volumetric light source. Composed of
several linear scales, each a different color, with density as transparency.
And respecting their true density when combined, so higher density scales
"shine through" lower. We'll see. Maybe overlay random sample "sparkles" to
maybe improve sense of spatial extent.

I've not seen much similar work. If you know of any, I'd love to hear of it.
Also, it seems there was a push some years back to emphasize electron density
when teaching intro chemistry. A push that doesn't seem to have gelled. I've
guesses, but I'd value thoughts on why.

~~~
natechols
I've been out of the field for a few years, but I know that the graphics
hardware (and accompanying software tools) make what you're describing much
easier. This is best example I can come up with:
[https://jcheminf.biomedcentral.com/track/pdf/10.1186/1758-29...](https://jcheminf.biomedcentral.com/track/pdf/10.1186/1758-2946-4-17)
(see page 12)

Random asides: 1\. once you're used to it, the simple isosurface mesh is super
easy to work with (i.e. build molecules into), and for structural biologists
the broad approximation is perfectly adequate 2\. even so, the amount of
unexplained blobs - some of which are definitely not noise - in my (xray)
electron density maps was always both fascinating and frustrating. that's one
reason why we don't spend much time on fancy renderings of experimental
density at anything worse than subatomic resolution, because you're just
sharpening the features that your atomic model doesn't explain.

~~~
mncharity
Thank you.

> once you're used to it, the simple isosurface mesh is super easy to work
> with

One wouldn't know it from the state of content, or even of much education
research, but educational representation is much harder than professional.
Students are unable to untangle features reflecting careful correctness, from
artistic license. So both seed mis/conceptions. Professionals can downregulate
misconceptions... though that can be surprisingly localized - asking first-
tier astronomy graduate students "What color is the Sun?"... gets you a common
misconception.

Consider that ball-and-stick wrapped with electron density in Fig 9. Imagine
coming at it cold. What is that stick? Well, maybe it's a ridge in electron
density, thus symbolizing the surrounding not-so-stick-shaped region of
increased density which constitutes a bond. But then what are those balls?
They're way too similarly sized to be electron density. Ok, maybe they're
spherical crosshairs for nucleus location, and the stick is a linear crosshair
for the ridge. Variously sized because... something. Oh, no, some of the
sticks are doubled, and density certainly doesn't have two ridges, or (here)
increased density. So we've a paper notation that badly misrepresents actual
electron distribution, blended into a physical representation with a... my
head hurts.

I saw a professional chem ed content discussion yesterday, around a
misconception, that in a two-species ionic solid, one atom bonds to another,
in pairs. Rather than to "4 to 6" neighboring atoms... because one common
printed diagram draws 4 neighbors, and another 6. It was like they were non-
scientists, thinking they could wordsmith their way to correctness, without
the slightest need to examine the actual characteristics of the real physical
systems being described, or to consult with someone deeply familiar with them.
The focus remained on models, decoupled from reality. I see a lot of that.

Chemistry education research describes chemistry education content using
adjectives like "incoherent". Maybe XR can serve as an excuse to do better?

~~~
natechols
All I can offer in response is vague, hand-wavy speculation about human
perceptive and cognitive limitations and how this impacts scientific
representations. Coming from a biology background, I'm very used to artistic
license and gross oversimplifications - and from years of habit, it almost
hurts my brain to think about molecules as anything other than chicken-wire!
From your complaint about those representations, I'm guessing you're
unfamiliar with "Richardson ribbons":
[https://en.wikipedia.org/wiki/Ribbon_diagram](https://en.wikipedia.org/wiki/Ribbon_diagram)

Even the developers of these visualizations would probably agree that it's
dangerous to rely on them too much, and they are only a way to convey specific
information in a way our eyes and brain can quickly process. Crystallographers
in particular tended to over-rely on those chicken wire views and that plus
software limitations yielded a lot of very poor-quality structures with poor
atomic packing. The developer of those ribbon diagrams (Jane Richardson) has
done a lot of other work to educate the field about how to visualize packing
and other molecular properties and avoid screwing up the analysis. Over the
long term, I think constant self-criticism makes up for the occasional
sloppiness in scientific research.

~~~
mncharity
My own focus is more on science _education_ representations. And on how badly
they describe the represented physical systems. Emphasizing that this seems as
much a science community failure, as an education community fail.

Consider an illustration of the solar system in some introductory astronomy
content. Let say, that not atypically, it misrepresents sizes, positions,
orientations, lighting, and Sun color. Thus creating and reinforcing
misconceptions known to be a problem in K-12, in undergrad, and even on into
astronomy graduate school. In contrast, consider a line representing an Earth-
Mars transfer orbit. It seems unlikely that students will say think there's a
material long pole there, and worry about it hitting satellites and cities.
Students and teachers are both clear on the aphysicality of the line, but not
of the Sun color. Hmm, though using a minimum-energy line to represent an
energy landscape _is_ a source of misconceptions.

Similarly, common representation of atoms and molecules are known to be
causatively associated with the stew of misconceptions that pervade students
and teachers of chemistry. Ribbons, perhaps not so much. Hmm, though also
similarly, they're often used without indication of regional flexibility, and
so perhaps contribute to the underappreciation of configuration landscapes, of
the importance of tuned floppiness. Perhaps.

Consider a currently implausible goal, of science education which accessibly
describes the physical world, and conveys a transferable understanding of it.
Arguably its content would look much more like scientific visualizations than
content does at present. But being for education, it faces additional
constraints, challenges and tradeoffs, distinct from those of professional
scientific visualization. Creating such would require a collaboration of both
deep scientific and educational expertise. For which very little incentive
exists at present.

I'm trying to come up with an XR-compatible visual representation of electron
density, that is physically correct, accessible, and bears in mind patterns of
misconceptions in chemistry education. That task should probably be more than
one random software dev's lockdown hobby hack. But as far as I know, that's
were we're at. The related NSF-funded work I've seen... doing this bit well
wasn't their focus. Same with XR ed tech side. Same with chem ed apps. And
scientific visualization programs. And chem ed research. There may well be
something nice out there, but I've not yet seen it. And big ed side... I was
chatting with a leading textbook publisher, which onboards content creators
with the indoctrination that their liberal arts background, and complete
unfamiliarity with science and tech, is not a problem... because there's "a
scientist" on call. Deriving electron density, with current nice python
libraries, and GPUs, has surely gotten vastly easier than it was with old
fortran messes, but still... it seems something more is needed here. Some
societal staffing seems missing. No?

------
presiozo
I thought an individual atom has been imaged before, at least roughly. Anyway,
this is fantastic science. I'm excited to see how imaging atoms affects our
understanding of quantum mechanics and atomic structure in general.

~~~
dmitrybrant
I think they mean that this is the first time individual atoms have been
imaged using this specific type of electron microscope. Individual atoms have
of course been imaged as early as the 1980s using scanning tunneling
microscopy. [1]

[1]
[https://en.wikipedia.org/wiki/Scanning_tunneling_microscope](https://en.wikipedia.org/wiki/Scanning_tunneling_microscope)

------
anfractuosity
I'm curious could you use AFM/STM to image molecules? I'm curious how that
would compare with this approach.

~~~
fabian2k
You can, and it has been done. But that was with very small, flat molecules.
Cryo EM can be used with very large molecules or complexes with many tens of
thousands of atoms.

~~~
jcims
Feels like this might be useful for exploring the surface of cancer cells to
identify receptors and other features that might be useful for developing
targeted therapies.

------
erik_landerholm
Dumb question, but if electrons are basically point particles, why is hard to
get below 1 angstrom resolution?

~~~
Areading314
At these scales, electrons behave more like waves than particles. Also the way
that they are scattered by the subject and manipulated via em lenses causes
information loss.

------
samovar_booth
For the record, individual atoms were imaged for the first time back in 1955:

[http://pubsapp.acs.org/cen/coverstory/83/8348atoms.html](http://pubsapp.acs.org/cen/coverstory/83/8348atoms.html)?

> On Oct. 11, 1955, Pennsylvania State University physics professor Erwin W.
> Müller and Kanwar Bahadur, who at the time was a Ph.D. student working with
> Müller, made history by being the first people to image individual atoms.
> The scientists were using a relatively simple and inexpensive instrument,
> and with it they directly observed individual tungsten atoms at the tip of a
> sharply pointed tungsten specimen.

------
jxramos
I had heard once that new imaging modalities that advance a lot of new
research invariably translate to Nobel prizes. Would that be expected of cryo
EM techniques?

------
The_rationalist
What would be the "realistic" potential use cases?

~~~
pritovido
We don't know yet. That is the idea about research.

When the microscope was invented, people made the same question: What is the
use of it? Just watching things we already know bigger?

That is a recurring question. So much that Aristotle created the book
MetaPhysics(meta meaning different from the Physics book) with the idea that
this particular book did not need to be useful to be created. It was a
compilation of non useful things.

This non useful compilation helped create science as we know it.

~~~
TomJansen
Oh please, this philosophical stuff is unnecessary. Cyro-EM is about a decade
old and is just an alternative to X-Ray Crystallography for elucidating the
structures of proteins. This article describes a way to improve the resolution
of Cyro-EM, matching it to X-Ray Crystallography. The biggest bottleneck of
X-Ray Crystallography is the production of protein crystals which is not
necessary with Cyro-EM.

~~~
twic
It's unbelievable how cryo-EM has come on. When i was a student, it was an
also-ran behind crystallography and NMR, something that a few weird groups
used to study a few special cases. Now you have to wonder if there's any point
doing crystallography at all!

I still like NMR, because you're looking at proteins in solution. But if the
freezing is good in cryo-EM, maybe even that becomes moot.

~~~
thereisnospork
I'm not sure if it applies to proteins, but for smaller molecules there seemed
to be some really interesting results a few years ago about using MOF's to
adsorb analytes in a spatially repeating manner so x-ray diffraction
techniques could be used without crystallization. Haven't followed closely
enough to know if anything is panning out though.

------
diegoperini
Where can I find the pictures?

~~~
COGlory
There aren't any. The way cryo-EM works is it images hundreds of thousands of
copies of the molecule and uses those to construct a 3D model, in this case,
with atomic resolution. However, a single micrograph doesn't necessarily
contain atomic resolution.

~~~
rezna
No longer need for hundreds of thousands. The aim is for just few thousands in
few hours to reach acceptable resolution.

The development goes real fast with both SW and HW.

~~~
COGlory
Depends greatly on whether or not what you're imaging has symmetry.

~~~
rezna
You can see that the record tests are being done with apoferitin or gaba. It
all depends on the improved qualiry of direct detection cameras, filters,
things like afis, fringe free imaging, ... The innovation goes insanely fast.

------
1-KB-OK
Makes me think of that one Simpsons scene where Mr Burns finds atoms in his
factory

[https://www.youtube.com/watch?v=DqgvHUg_vxY](https://www.youtube.com/watch?v=DqgvHUg_vxY)

------
Gatsky
Looks like atomic rez is only possible with certain very stable proteins.
Hopefully the tech will improve to allow broader application.

------
auvi
Now I would like to see a Corona virus with it.

~~~
sixstringtheory
Another commenter linked to a study that did just that:
[https://news.ycombinator.com/item?id=23415625](https://news.ycombinator.com/item?id=23415625)

------
yters
how do we know atoms exist?

