I'm writing this blog mainly for the students at Peterson School so that they can get a sense of what doing science is like from a Peterson alumnus (me) who is now a scientist in training.
Hi Peterson students and friends! I've arrived at the University of Cambridge and started working on my new research project, which involves doing computer simulations of protein folding. I'm doing this work with Dr. Robert Best, a fellow in the Cambridge Department of Chemistry. I'd love to tell you all about it and answer any questions you might have.
This will hopefully be the first of a series of posts on my blog about what proteins are, how they fold into complicated 3D structures, and why that is important for life. I apologize in advance if some of what I'm saying is unclear. It will hopefully become clearer as I go on...
But first, here's an animation of one of my first simulations. The coiled tube represents an amino acid chain (a.k.a. polypeptide). You can see three major coiled parts of the chain that pack against each other. These are called alpha helices--more on those later. The structural motif that they form is called a three-helix bundle. The animation shows what happens to the simulated protein if you add to the simulation a strong pulling force between the two ends of the polypeptide chain. The protein quickly comes apart or unfolds. (People have actually done this sort of experiment in real life...but more on that later!)
Asaad asks a really good question about the safety of sticking random genes into people. To be honest, I can’t give you a definite answer about whether sticking GFP into a human would be harmful or not because, as far as I know, it has never been tried. One major ethical concern about genetic engineering in humans is that we can’t be 100% certain what sticking a gene into a person would do until we actually do the experiment. This is less of a problem for conventional pharmaceuticals, because you can recruit volunteers who are informed about the risks. But a person who isn’t even born yet obviously isn’t around to make that sort of decision! A person who has been genetically engineered would be stuck with that for the rest of his or her life. And you are right that the children of that person could inherit the gene. (They would probably have a 50% chance of inheriting it if it was carried on one of the parent’s chromosomes.) Whether or not it would ever be justified to make that sort of decision for someone is a very serious ethical question!All that said, though, my guess is that GFP would probably be safe to have floating around inside your cells. As far as we can tell, it doesn’t seem to do much of anything (aside from glowing) when you stick it into the cells of other organisms. So there isn’t any reason to think that it would be harmful…though it probably isn’t work risking it just for fun!As for how fluorescence works…that’s a really cool story. You may have talked a bit about atoms in one of your science classes. If you remember, atoms are made of nuclei (made of positively-charged protons and uncharged neutrons) orbited by negatively-charged electrons. The electrons are in things called orbitals. You can think of an orbital as like a “cloud” of electrons with a particular shape. (If you study quantum mechanics someday, you’ll learn that electrons in orbitals can be thought of as waves too…that’s a weird thing to think about!). Well anyway, atoms can combine in different ways to form molecules. When this happens, the electron orbitals from all the individual atoms fuse into new orbitals called molecular orbitals. When the molecule is just hanging out (i.e. when the light is off), those electrons are generally in the lowest-energy orbitals possible. However, when a particle of light (a photon) with the right energy interacts with the electrons, they can absorb the energy in the photon and jump up to a higher-energy orbital. The molecule is in what is called an excited state. However, this excited state is unstable. Think of it as sort of like a pencil balanced on its tip. Maybe the molecule can exist in that state for a split second, but it wants to get back to its ground state. In some molecules, the electron just falls back to its low-energy state and the energy from the light is lost as heat. But in some special molecules (like the molecules of the fluorescein that I brought you), something really cool happens where the molecule emits a new particle of light when the electron falls from its excited state back down to its low-energy state. This is the light that you observe as fluorescence. The new photon of light is a different color than the original photon of light because some of the energy is typically lost in the process of kicking the electron up to a high-energy state and letting it fall back down again. I hope that wasn’t too unclear! Let me know if you have any more questions! Note: 1 nanosecond = 1/1,000,000,000 second 1 picosecond = 1/1,000,000,000,000 second 1 femtosecond = 1/1,000,000,000,000,000 second!!!!!!
Thanks so much for letting me give a presentation for you guys! I had a lot of fun, and I enjoyed meeting all of you. I want you to know that I was incredibly impressed with what you knew already, and I was even more impressed by the sorts of questions that you asked. Being able to ask good questions is one of the hallmarks of a good scientist. If you have any more questions that I didn’t get to answer, please post them as comments on my blog, and I would love to try to answer them. (And if I don’t know the answer, I will try my best to find out.) Also, good luck to those of you who are going to the science fair! I’m sure you’ll do great.
I will keep posting new stuff on my blog (and I’ll try to get friends of mine at UChicago to post stuff as well), so I hope all of you will keep in touch. If you have any suggestions, feel free to add comments.
Thanks so much for letting me give a presentation for you guys! I had a lot of fun, and I enjoyed meeting all of you. I want you to know that I was incredibly impressed with what you knew already, and I was even more impressed by the sorts of questions that you asked. Being able to ask good questions is one of the hallmarks of a good scientist. If you have any more questions that I didn’t get to answer, please post them as comments on my blog, and I would love to try to answer them. (And if I don’t know the answer, I will try my best to find out.) Also, good luck to those of you who are going to the science fair! I’m sure you’ll do great.
I will keep posting new stuff on my blog (and I’ll try to get friends of mine at UChicago to post stuff as well), so I hope all of you will keep in touch. If you have any suggestions, feel free to add comments.
One of my favorite experiments to do in my lab involves shooting cells with lasers. Yeah, that’s right…shooting cells with lasers!It’s part of a really cool procedure called fluorescence recovery after photobleaching or FRAP for short.
The fluorescent proteins that I wrote about before are useful way beyond just making glowing fish.They can also be fused to other proteins.This is done at the DNA level by taking the gene for one protein and fusing it to the gene for a fluorescent protein.The molecular machinery inside the cell transcribes this DNA onto a single RNA which is then translated into a single protein called a fusion protein.Fusion proteins can often do the same thing that the original protein did.(It’s very important to test that this is the case.Sometimes tacking on an extra piece can disrupt the protein’s function!)Here’s a picture of a fusion protein that I made. The protein is in the nucleus of a living cell, as seen under a fluorescence microscope.I have done various tests to make sure that it still does what the normal protein does.So far, it has passed all of my tests in cultured cells (in a Petri dish, basically), and I am now working to do similar tests in living flies.
So, what is FRAP?FRAP is a technique that can be used to study how fast fluorescently tagged proteins move around inside cells.The idea is actually pretty simple:You take a laser and zap the fluorescently-labeled protein in a little spot.(You do all of this with a high-tech microscope.)The laser is so intense that the fluorescent label goes dark or photobleaches.This is sort of like what happens when you leave colored construction paper out in the light for a long time.The dye molecules in the paper undergo photochemical reactions (chemical reactions caused by light) that cause them to lose their color.Similarly, a fluorescent molecule photobleaches when it undergoes a photochemical reaction that turns it permanently into something non-fluorescent.
The rest of the fluorescently-labeled protein, however, is still okay.It still glows as before.If the protein is free to move around, then the bleached protein inside the spot will diffuse out and the fluorescent protein surrounding the spot will diffuse in.You can determine how mobile a fluorescent fusion protein is by measuring how long it takes for this exchange to occur.The result is a recovery curve, like the one below.Proteins that are free-floating in the cell recovery quickly after photobleaching.Proteins that are stuck to larger things recover more slowly.You can figure out a lot about what a protein is doing in a cell by comparing FRAP curves of different mutants under different conditions.
In addition to FRAP, there are even more advanced fluorescence techniques that you can use to study how proteins behave in living cells…more on those later.
I'm an masters (MPhil) student in Chemistry at the University of Cambridge. I'm working with Dr. Robert Best to investigate using simplified models how the folding pathways of proteins are influenced by mechanical forces and chemical denaturants. Next year, I will return to the United States to do a Ph.D. in Systems Biology at Harvard University.
