Biochemistry Question of the Day: 19 April

Isn't 'the destruction box' the greatest biochemistry term ever? Any challengers?

A very well written piece

Blake Stacey wrote a great piece outlining some of the reasons why rational people get upset about lies and hate spread by certain groups. It's actually kind of scary, and more than a little depressing, but it's well written, and I think it's something more people should be aware of. Go check it out, and maybe after you've finished reading you can teach your son, daughter, niece, nephew, husband, wife, parents, siblings, friends or anybody with whom you happen to be talking something about the wonders of science.

How will I be able to judge it myself?

Apparently Expelled won't be coming to Canada until this summer. Sigh.

Protein Crystallography: How?

Well, I said about two weeks ago that I would do another post on crystallography. And then I didn't. But I will now!

So I've talked about why we want to crystallize a protein, now I'd like to tell you about some techniques we use to actually do it!

In general, a crystal isn't a natural form for a protein to be in. (In fact, in a lot of cases, proteins aggregating together can be a big problem.) So it's not a simple thing to make them crystallize. It can be downright frustratingly impossibly.

To make a crystal, you need to have individual molecules come out of the bulk solution. The trick is that when you take a protein (or really any other molecule I guess) out of solution, it'll probably just form an ugly precipitate. An ugly precipitate is useless for crystallization as there is no regular structure.

So the general method is to cause it to very SLOWLY come out of solution, in the hope that as it does, it'll stack in that lovely regular lattice pattern. To make it come out of solution, the solution must be saturated, that is, it must have as much protein as can possibly stay dissolved in the particular solution. Then, you make it a little more concentrated, so it'll be more than saturated. Some will have to come out.

The way I know to do this is called vapour diffusion. The concept is pretty simple, you have your saturated solution, and then you let some of the water in the solution evaporate out, causing the solution to be slightly more concentrated, hopefully enough for the protein to crystallize out, but not so much that it precipitates. Commonly, a drop of your protein solution is placed in a closed environment with some hygroscopic (water drawing) substance. Two common setups are hanging drop and sitting drop vapour diffusion:

On the left is my representation of a hanging drop setup. A well is filled with the hygroscopic solution, and a drop of protein solution is placed on a coverslip and hanged over top the well solution. (The well must be sealed from air of course) The sitting drop setup is very similar, but instead of hanging the drop from a coverslip, you sit it on a pedestal above the solution. I guess the names kind of gave it away. In either case, the solution in the well will suck some water out of the drop, and you'll look through your microscope and hopefully find some tiny little crystals. Eventually. This can take days, weeks, months to actually produce crystals.

This would also usually be done on a plate containing many wells (24, 96, etc.), so that a range of different conditions could be tested at the same time.

Now, once you've grown your crystals, that isn't the end of the game. You have to somehow manage to mount a single crystal so that you can shine your beam of RADIATION through it and create that diffraction pattern I mentioned and THEN do a load of work to construct an actual idea of a 3D structure.

Still, you'll be pretty excited once those crystals first show up.

There's actually other methods used as well, but seeing as I don't know anything about them it's probably best for me to just stop here. Did this all make sense?

Science TV?

I don't watch any real television these days. I have a tv set, but all it's hooked up to is a vcr, dvd player, and super nintendo. Now I'm finishing up school pretty soon, so I might have the time (and money) to watch a little bit.

What I'm hoping for is a good science program. Something like Bill Nye the Science Guy, or Cosmos. Is there anything out there these days? Any favourites of yours? Or should I just buy Cosmos on DVD?

It All Comes Together

So a few days after I complain about not hearing very much about the third domain of life, I'm required to write a paper about some archaea! Well, actually a protein from archaea. It also links to my post about crystallography, as this was the first protein to have it's structure determined to atomic resolution using a related crystallographic technique: electron crystallography!

So, Biochemistry Question of the Day:

Name the protein.

Protein Crystallography: Why?

Tonight, in honour of my first presentation of research I've done myself (as small as it was) I'm going to spread some information. I think the average person doesn't know anything about protein crystallography, and most scientists, even biochemists, only know that it exists and what it gives them, but don't know many of the details.

I'm not an expert in this of course, but I know more than the average person, so here goes:

Proteins are very small. Incredibly small. Now they vary in size of course, from a few amino acids (some people would call these peptides, same basic structure though) to many many more amino acids, still we basically can't see any individual protein. Not only can we not see them with the invisible eye, we can't see them with light at all. I can't see cells with my eye, but if I put a cell under a microscope, it's big enough that I can still see it. Proteins are so much smaller than that (obviously, since cells have thousands of proteins in them). We can't see a protein molecule even with a light microscope. They are smaller than the wavelength of the light we see.

But we want to see them. Proteins are key to life, and understanding how life works is what science is about. If we could see proteins, it would give us so much information, about how they work, how they're built, how they interact, how they go wrong, etc. etc. So how do you see something that is too small to see with light?

Use x-rays! X-rays have wavelengths shorter than the visible light we see, so we should be able to use them to see these tiny things: molecules like proteins.

Problem! When we use a light microscope, we use lenses to focus the light to make an image we can see. There's no lens for x-rays. Dang, does that mean that using x-rays to see proteins is right out? Well no, because of some very clever. When you shine an x-ray source on a molecule, it'll be deflected in a regular pattern, and if you're really clever, you can use that pattern to build back a picture of the molecule.

This brings up another problem. If you shine your x-rays on the single molecule, you won't detect enough deflections to actually be able to put a picture together.

Okay, keep that in mind. We're going to talk about crystals for a second.

We all know about crystals. Salt is little crystals. Sugar is little crystals. When I was a kid I grew some larger crystals of salt, you can easily do that yourself. But what exactly are they? Crystals are a structure made of a repeating pattern of. . . something. Atoms, ions, molecules.



This is what a salt crystal might look like (in two dimensions, imagine those are all actually sphere shaped, and the pattern has depth as well). This pattern will repeat until the ions run out, which would create the edge of the salt crystal.

So imagine we could do this with a protein molecules, get them into this regular lattice that repeats, in the same orientation in space over and over again. Then you could shine your x-rays on it like before, but now the deflection pattern would be strong enough to be useful, because each deflection would be repeated over and over again. With this deflection pattern, we could build up the model of our protein. Awesome!

This works. In fact, anytime you've even seen a structure of a protein, it was probably determined from x-ray crystallography. It's been done thousands of times, in the last forty years. The first structure that was solved was forty years ago. It was myoglobin, and from what I've read, it was really exciting. In fact, the people who did it got a Nobel prize for it in 1962.

I hope this made sense, and also that I got the facts right. If not, come yell at me and I'll fix it. For more, see: How we crystallize proteins!