Myoglobin as a Probe for Understanding Molecular Evolution
Primary Author: Ajit Chary,
Undergraduate Research Assistant
IntroductionFirst of all, what is myoglobin? What's your favorite sport? Let's say basketball. Now you need a lot of strength to play basketball well, right? And what gives you the strength to jump up and dunk the ball like Jordan? That's right, MUSCLES.
Now, how does a muscle get big? Well, you can lift weights, and you can jog, run, jump, and play outside everyday. That's all good. But then you need something to give muscles enough energy to let you jump, lift weights, play, jog, run and dunk. What lets the muscle get big and strong and what lets the muscle live is oxygen, found in the air you breathe.
But how can we breathe in through our noses and get oxygen to our arms, legs, and all the rest of our muscles? The way the oxygen gets there is through the blood. Then, the blood goes through our muscles and gives it the oxygen.
Where exactly where does the oxygen go when this liquidy red stuff gives it to the muscles? And how can the oxygen stay in the muscles so that we can use them and get energy from them? Through a tiny little protein called MYOGLOBIN.
So that means that every single thing that has muscles, meaning all living animals and people have myoglobin right? Therefore is it all the same? I mean does everyone and everything, even your pet dog, cat, or a whale have the same myoglobin as we do? If not, then is it similar to ours? Lets find out.
Before we take a look at the Biology Workbench, we suggest that you either print out a copy of this tutorial or open another web browser window to view this tutorial while playing with the Biology Workbench.
Using the Biology WorkBench
The first item you will see is a gray query box with several parameters. On the right side of the search bar, you will see "Show 10 Hits". Change that (by clicking on the icon) to "Show All Hits".
What BLASTP does is quite interesting. It takes the human that you selected and compares it to that huge protein database (for now let's make it PIR) or another huge database (like SWISSPROT). You are permitted to select one or more databases to compare. Now since you clicked human, BLASTP will find all of the animals, plants, even bacteria and viruses, that are similar or the same as the human protein you selected (in this case myoglobin). It doesn't stop there, it also does a big favor by putting the protein sequences in ORDER for you, so you can see what is the closest (evolutionarily) to humans and what is furthest from humans. Since myoglobin is found in muscles and animals with blood, you won't see any plants or viruses. But, if you use other sorts of proteins like Enolase, or Aquaporins (higher level proteins) you can and probably will find all of that. What animals do YOU think have myoglobin thats the most like the myoglobin that we humans have? Dogs? Cats? Whales? Monkeys and other primates? Birds?
If you guessed Primates, you were right. Now, which PRIMATE is most closely related to humans? Lets see...
DO NOT CLICK ON HUMAN because you already are comparing everything to a human. After you get used to this BLASTP program, choose any animal you want and see how closely related it is to humans. After you clicked these animals, hit Import Sequences.
Clustal W is a program that will let you see the sequences and it will align all of the sequences for you so that you can see for yourself how closely related and how far apart all they are. In this case, it will show you how closely related they are. These sequences are shown in letters. These letters represent chains called Amino Acids which all form together to make the Myoglobin protein structure. The letters are compared with other letters, and you will see that the letters are arranged columnwise, and that several of these letters (by column) are the same. Don't really worry about what all these letters mean. At the beginning of the page, hit "Import Alignments."
After you hit this, you will see a little gray box and to the right of the box, you will read CLUSTAL-W Protein -- and all of the sequences you picked.
Click this box, and then look at the large scrollbox above it.
Scroll down the box with the programs within it, until you see
Now, at the begining of this new page hit submit, because all of the default parameters work well.
After you take a look at the flowchart, hit the Return button in the Biology Workbench. That will take you back to the place where you selected the DRAWGRAM and DRAWTREE programs. Now repeat the procedure with DRAWTREE, view the unrooted tree and hit "Return" once again.
Now to compare the sequences with another program, you can go to BOXShade - Color-coded Plots of Pre-Aligned Sequences, and on the next page hit Submit.
Now take a look at the COLORS. These colors have a relationship and compare the sequences in this manner:
Now after exiting BOXShade by pressing Return, you can start the wahole process again with a different organism, different BLASTP database, (like SWISSPROT), different set of animal comparisons, and you can play around with even different proteins that you know of.