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.