This past week in lab will be known as “The week I used a lot of radioactivity.” I didn’t enjoy it. I had to get dolled up in my lab coat (yup, found it) bearing my dosimeter, another dosimeter on my right index finger, two layers of gloves, and some sweet yellow safety glasses. Not part of the official outfit, but necessary nonetheless, is having long hair tied up, closed toe shoes, and long pants. During an unexpectedly warm day a few weeks ago, I had this attire on and our lab was a sweltering 90 degrees. I was an unhappy post-doc.
So, why in the world was I playing with radioactivity all week?
Scientists use it quite often, actually. Aside from the safety headaches, radioactivity is incredibly useful to understanding exactly what is going on with our microscopic little proteins. To explain how, I’m going to use a technique that I often use with my chemistry tutorees or younger labmates: You have to crawl inside the experiment.
Let’s go.
Experiment: You have two tubes. One tube contains water. The other has a protein floating around in it. Which tube is which?
More Useful Information: The protein is a particular kind of protein known as a kinase, which is an enzyme. Enzymes are very specialized and typically just do one type of job. Think of a factory where cupcakes are being made (is the analogy getting old yet?): one person makes the batter, one person fills the cups, one person bakes them, one person ices them, etc… Each person in that line is doing a very specific job. In our cells, enzymes are much like those cupcake assembly line workers. Each enzyme has its own small task in the great scheme of a larger process. There are thousands of jobs to do so there are a lot of enzymes. Kinases are a particular class of enzyme. The job of all kinases is to move a phosphate group from ATP to another protein (target protein) (Figure 12.1).
So… good. What do we do now?
Plan 1: Let’s add a little ATP and target protein to each tube. The tube with the kinase will then have target proteins covered with phosphate groups (Figure 12.2).
Problems with Plan 1: How can we tell if the target protein is covered with phosphate groups? It’s not like we see the target protein.
Solutions to Problems: While there are ways that scientists can “see” proteins, the easiest experiment to do here is to use a little radioactivity*. 32P to be exact.
What is 32P? Phosphourous is element 15 on the periodic table. A particular type of phosphorous has 17 neutrons and 15 protons (15+17 = 32) in its nucleus. This nucleus is unstable and will decay very predictably. We can purchase ATP with a 32P incorporated such that when the kinase moves the phosphate group, it will take the 32P (Figure 12.3).
I’d like to take a moment and stress here that the phosphorous in our bodies is not radioactive. Much like gloves in different colors, elements can be found in different forms. Some of the forms are radioactive, but most are not. Since ATP contains phosphorous anyway, scientists can switch out a nonradioactive phosphorous for a radioactive one and sell the crazy molecule to post-docs like me to do experiments.
Plan 2: Add a little radioactive ATP and target protein to each tube. In the tube with kinase, the target protein will become radioactive. The target protein in the water tube will not be radioactive.
Problems with Plan 2: How do we separate the target protein from everything else? How can we tell if it is radioactive?
Solutions to Problems: Special paper exists that will bind protein molecules but not ATP. Special machines will tell you if that paper is radioactive or not.
Excellent! So off we go to do our experiment and the results are in Figure 12.4.
Which tube had the kinase and which tube had water?
Think about the caption of Figure 12.4, as well. Is what is bound to the paper makes sense given everything you’ve read above? Have you successfully “crawled inside the experiment?”
Experiments like this are done very often by scientists. Obviously they are asking more interesting questions than which tube holds the kinase, but the basic experimental set up is what I outlined above. These are the types of experiments I was doing this week. Unfortunately, I think that next week will be called “The week I used even more radioactivity.”
Fun Facts that I couldn’t fit in the post:
1. I’m sure most people know that radioactivity is unhealthy. One reason why is because the energy radioactive decay emits can cause mutations in our DNA. Ooooh… think about the last post (Cancerous Mutational Problems).
2. All work with 32P, which is known as a beta emitter, must be done behind a plexi-glass shield. It can be super awkward wrapping your arms around the shields to do your experiments.
3. Radioactive ATP comes from Perkin-Elmer in little containers that block the radioactive decay. They are quite cute.
4. Radioactive ATP is colored lime green. (I assume so that you can see it if you spill some.)
5. Scientists wear dosimeters so the powers that be can know how much radioactivity we are being exposed to. Using proper shielding should result is very little exposure, but they check the dosimeters to be sure we are working safely. I had a friend who hung his coat (plus dosimeter) by the radioactive waste one day. When checked, his dosimeter was off the charts. Yeah, they were certain he had cancer until he explained.
6. Purchasing radioactivity is incredibly regulated. 32P represents only one of hundreds of different products and one of several different kinds of available radioactive elements for laboratory use. Some common other ones: 3H, 35S, and 125I.
7. Scientists follow very strict guidelines on radioactive waste disposal. Most is safely stored until a large proportion of it has decayed. We certainly do not pour it down the sink or drop it in the trashcan when we’re finished!
* Any scientists reading this are probably laughing. I realize this is like using a hammer a push in a thumbtack, but I’m using a simple and straightforward experiment to illustrate a point. Go with it. If this was real life, I’d use Bradford Reagent and know in five seconds. But this post isn’t about Bradford Reagent. Maybe I’ll do that one another day!
Dosimeter: a small plastic device (~ 2 inch square) that records how much radiation one has been exposed to while working with radioactivity.
Enzyme: a protein that catalyzes specific reactions
Kinase: a specific class of enzymes that transfer phosphate groups from ATP to target proteins
Target protein: the protein which receives a phosphate group in a reaction catalyzed by a kinase
Bradford Reagent: a red dye that turns blue in the presence of protein
REFERENCES
Me, myself, and I.
Perkin-Elmer: www.perkinelmer.com
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