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Cryo-electron microscopy breaks a key barrier

nature.com

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Re: Cryo-electron microscopy breaks a key barrier

#101

Post by mensetmanusman »

COGlory wrote: post #39

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 wo…

Can you use CryoEM to see the atomic bonding arrangement in any material you throw at it? E.g. visualize a glass or polymer network?

Re: Cryo-electron microscopy breaks a key barrier

#102

Post by [deleted] »

fabian2k wrote: post #11

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…

[deleted]

Re: Cryo-electron microscopy breaks a key barrier

#103

Post by Koshkin »

hoseja wrote: post #96
m4rtink wrote: post #88

Earlier quoted context omitted.

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 t…

Small world :)

Until you start counting them in moles.

Re: Cryo-electron microscopy breaks a key barrier

#104

Post by Koshkin »

voxelghost wrote: post #90
mysterypie wrote: post #65

Earlier quoted context omitted.

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…

The nucleus is on the scale of 10e-5 (1/100,000) to the size of the whole atom (cloud).

Yet, the maximum density of the electron cloud (at least in the case of s-orbitals) is right inside it. (Talk about the "size of the whole atom"!)

Re: Cryo-electron microscopy breaks a key barrier

#105

Post by dekhn »

natechols wrote: post #48
fabian2k wrote: post #11

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…

When I started grad school in 2003 there were a handful of structures approaching 0.4 nm, which is the point where you might be able to build most of the sequence into the map de novo. But those were exceptional cases.

my favorite report from grad school was the paper that had to be retracted because they "built the sequence into the map backwards" (IE, the N and C termini were switched).

Re: Cryo-electron microscopy breaks a key barrier

#106

Post by natechols »

dekhn wrote: post #105
natechols wrote: post #48

Earlier quoted context omitted.

When I started grad school in 2003 there were a handful of structures approaching 0.4 nm, which is the point where you might be able to build most of the sequence into the map de novo. But those were exceptional cases.

my favorite report from grad school was the paper that had to be retracted because they "built the sequence into the map backwards" (IE, the N and C termini were switched).

Was that the Great Pentaretraction or something else? My favorite is still the Krishna Murthy structures that turned out to be completely made up.

Re: Cryo-electron microscopy breaks a key barrier

#107

Post by rezna »

COGlory wrote: post #72
rezna wrote: post #60

Earlier quoted context omitted.

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.

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

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.

Re: Cryo-electron microscopy breaks a key barrier

#108

Post by rezna »

m4rtink wrote: post #88
jjoonathan wrote: post #19

Earlier quoted context omitted.

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?

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 t…

I work with latest greatest Krios machines, and all the latest HW and I make SW for it. It's great job.

Re: Cryo-electron microscopy breaks a key barrier

#109

Post by jgehrcke »

Ultimatt wrote: post #97
jgehrcke wrote: post #76

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…

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 c…

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.

Re: Cryo-electron microscopy breaks a key barrier

#110

Post by COGlory »

natechols wrote: post #100
COGlory wrote: post #80

Earlier quoted context omitted.

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 ne…

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 approa…

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.
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