Understanding Crystallography – Part 1: From Proteins to Crystals

Understanding Crystallography – Part 1: From Proteins to Crystals

This is a three
dimensional structure of a protein called lysozyme. It represents the
order of nature. And I feel awed
when I look at it. It’s an enzyme that’s in our
tears and saliva and mucus, and it helps us fight bacteria. And knowing this three
dimensional structure helps us to understand
the mechanism of action of this enzyme,
and therefore how it helps fight
bacteria in our bodies. Unfortunately, the
size of the molecule is such that it’s
far too small for us to see under a light microscope,
or with our naked eye, because the wavelength of
light is much, much larger than the size of
this tiny molecule. For that reason, we
have to use radiation that’s much smaller
in wavelength. And we use X-rays to
look at these molecules. These are protein crystals. Locked within each crystal
are millions of protein molecules, all arranged in an
ordered, grid-like structure. By firing X-rays at these, and
measuring how they scatter, we can work out the
molecular structure of nearly any
crystallised sample. It’s through this method,
known as X-ray crystallography, that some of the most
important biological structures have been obtained. from the double helix of DNA
to numerous proteins, vitamins, and drugs. But getting from a
crystal to something like this, the structure,
is not at all trivial. And it can take a long time
to grow a suitable crystal. Now we’re in the Protein
Production and Purification lab, because before we can
set up crystallisation trials, we need to produce enough
protein for that process. We do this by genetically
modifying E. coli bacteria, which then acts as
a little factory to produce our protein in
a large flasks of broth, which we incubate. Once that’s happened,
we break open the cells, extract the protein,
and then purify it, ready for the
crystallisation process. So why do we need to grow
crystals of our protein molecules before we can
shine X-rays at them and try and find the three
dimensional structure of them? Well, if we imagine that this
string of beads is a protein molecule made up of 20
different amino acids– different colour of beads–
that are found in nature, this folds up in a very complex,
complicated manner in the three dimensional shape, like this. And if we look at one of these–
a true biological molecule here, a real one– what
this metal model represents is a string through the beads. And you can see it
starts at this end, and it follows a pretty
torturous path going around here, that you’d never
imagine when you just look at the string
of amino acids. And the other end of
it comes out here. Now it turns out that
if I take a tube here with my protein in it, that
I’ve purified and prepared, there’s millions and millions
of protein molecules in there. And if I shine X-rays
at it, the X-rays will scatter off in all
sorts of random directions. And I won’t get any
information about the shape of the molecule within the tube. However, if I can get the
protein molecules to line up in an ordered array–
such as in a crystal, where they’re all lined up in
the same orientation– when the X-rays scatter
from the crystal, then I can get
enough information. The signal is strong enough for
me to get the three dimensional structure of the protein. We’ve now come down to
the crystallisation lab to look at how we
crystallise proteins. In this Petri dish, I’ve got
some supersaturated sodium acetate. And that means that there’s
so many molecules crowded in this solution, it’s almost
not holding the molecules. And it wants to solidify. And if I hit it
with a spatula here, you can see that
it crystallises. We get a fantastic pattern as
it crystallises across the dish. Essentially, this is what we
try to do with our proteins. Which is to produce a
supersaturated solution of the protein, and we dehydrate
in a very controlled manner. The proteins we work
on here unfortunately can be sometimes really
difficult to crystallise. So we load small volumes of the
protein into trays like this, with different additives. But we have robots
that help us do that by pipetting small
volumes into these trays. Once we have the tray with
the protein and additives in, we take it around to
this crystal hotel which holds the tray for several weeks
at four degrees centigrade. And also monitors whether
we have crystals or not by photographing the
tray drops regularly. But that’s only part
one of the story because once we finally manage
to grow a protein crystal, we then have to take
it for X-ray analysis. And from the data
we obtain, we try to generate a structure
of our protein molecule, such as this one of lysozyme. But the protein structures
we work on today are far more complex. And they can produce very
small and delicate crystals. So to study them,
we have to take them to extremely powerful
X-ray sources at specialist facilities, such
as the Diamond Light Source. It’s only once we get
our crystals there that the next stage of our
journey can truly begin. It’s the most expensive
and sophisticated scientific facility
ever built in the UK. The instrument in
this building can produce X-ray beams
powerful enough to peer right into the atomic
heart of all kinds of matter.

51 Replies to “Understanding Crystallography – Part 1: From Proteins to Crystals”

  1. How can you determine the structure of a complex molecule too small to see under a microscope? This video will make things crystal clear… 

  2. Around 2:55, where the little crystals grow out of the blob of liquid, is that an actual recording of the process under a microscope, or is it just nice CGI?  'cause damn, it looks awesome!

  3. I'm yet to see a biochemist discuss how much of the crystal structured determined is actually relevant in biologic medium.

  4. This is rediculous. You people are mental. Human beings truely are amazing creatures. What's the name of the lady Watson and Crick stole this idea from?

  5. I'd really like to know what the real application of the knowledge about 3-dimentional molecule structure is. Can't figure this out from the information on crystallography

  6. If possible, I for one, would be really interested to know exactly how complex components (the stuff that has been put in layman terms) work

  7. I think one of the reasons I find this all fascinating is because I am NOT taking a class on it, I just watch it out of curiosity, I am sure if I was going to be tested on it I would be far more frustrated by the material.  My suggestion to anyone is always learn as much as you can on your own time so that when it comes down to being required to know or learn something you will already have some inkling of knowledge of the subject matter and it will still be somewhat fun but not nearly as daunting as if you were witnessing it for the first time blindly shrouded in the pressure of needing to learn it all before a a major exam.

  8. This video was very educational and also interesting. Usually these types of videos don't help but this one was really helpful.

  9. Could you please explain me the basics of why the detection limit of a photon is strictly associated with its wavelength? Why a "visible" photon is not good per se to get info at the atomic level. Just to make an example infrared spectoscopy can be used to study molecules whose size is far below IR wavelength. Sorry for being naive 🙂

  10. Strictly speaking, one can indeed get an idea about shape of molecule by measuring scattering of protein in solution. Called SAXS…

  11. wow the filming of that was absolutely terrible. just in and out of focus. always zoomed in. no actual view of the entire structure. we as viewers have no idea how large or small it is.

  12. Thank you very much, I now has a hope for accomplishing my structural bioinformatics class, which kind of frightened me today! (But it's still just a hope only.)

  13. This was so cool! thank you. PRobably one of the neatest and most informational productions I've watched to aid in my learning.

  14. Now this is a video. I'm a chemist and have taken graduate quantum chemistry and organic chemistry courses at UVA. All my life I'm wondered how anyone can "get into" either chemical group theory or crystallography. If I'd seen this demonstration, I might have pursued both.

  15. this is amazing! I only learned little about it in one of my bio lectures so it's good to have a better understanding of it because it is actually so interesting !!

  16. I learned about X ray crystallography in my physics class when we went over interference and diffraction. Just searched for relevant videos on youtube, and I find out it has this crazy biological application! Science is such a trip lol

  17. So uh this video is quite old but i was wondering if there's anyway for me to receive more in depth information about all of this especially the actual process behind crytallising and what these additives do to the protein. If yes please do reply and thank you very much this video was more informative than hours of google.

  18. I find it so fascinating how minute changes in structure can have such a large impact on a compound’s properties.

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