Alarm goes off: it’s time to wake up… already?
I recently started to walk to work, a healthy and cheap alternative to the packed Northern line trains. I quite like changing route every day. I often get lost, but the Shard always shows me the right way to Guy’s Campus.
It’s time to start my experiments. In my PhD I am studying the mechanism of action of a human enzyme involved in the regulation of blood pressure. For all my Biophysics experiments I use loads of protein, which means I cannot isolate it from human tissues. This is why I produce the enzyme using bacteria engineered to generate it. Yes, you got it right: I have bugs which I command to produce the protein I study. Doesn’t it sound cool? Well, it does to a geek like me…
First thing to do is to start the bacteria cultures and, while they will grow and produce the protein for me, I can carry on with the rest of the experiments.
Last week I cultured some cells which had already produced the protein, so I can extract and purify it. To do this, I break the cells so that the protein goes in solution. Then using different resins which specifically bind the protein, I remove all the contaminants and get a super pure sample. Nowadays purifications are made easier by using automated systems such as the one in the picture, called AKTA.
While the AKTA purifies the protein I can head to lunch!
It is finally food time. I usually have lunch with some other Randall pals. We are always the loudest table: sometimes embarrassing, but definitely fun!
No more experiments can be done without an adequate dose of caffeine. It’s coffee time.
Strictly espresso by the way (I’m Italian after all).
It is time to get back to work and find out if the protein purification worked. I will run an SDS PAGE, an analytical method to evaluate the purity of the sample obtained. We biophysicists are quite crazy for purity: either is super clean or we start again!
Happy Days: I obtained only one band, which means the protein is highly pure. I can go on with the rest of the planned experiments.
Very often here at the Randall we have international guests presenting their work and scientific achievements. Today’s speaker is coming from Germany and will show us his studies carried out using a technique called Cryo-electron microscopy. It is one of the most cutting-edge technique in Biophysics field and I am really excited about learning something more about it.
The seminar was definitely inspiring, but it is time to go back to the lab.
One of the goals of my research is to discover novel drugs to target the protein I am studying. Indeed, this enzyme induces hypertension in several cardiovascular diseases and this is why I am looking for molecules able to inhibit it. Today I will test one of the compounds of my library using a spectrophotometric assay.
The curve looks pretty awesome! The wavy shape, called sigmoidal, suggests that the compound I have tested can inhibit the enzymatic activity. I found a new drug candidate for my studies. But do not get too excited: I still have to find out how the compound binds to the protein. To do this I perform X-ray crystallography experiments, a sort of hypersensitive photographic technique that allows you to take pictures of proteins.
Speaking of which, I have to check my crystals!
Last week I set up crystallisation trials. In this experiment the protein packs in very ordered microscopic crystals, which can be shot with X-rays. According to how the light diffracts, it is possible to solve the structure of the protein and have its picture.
Awesome, the crystals grew! I will fish them with a microscopic loop and soak them with the compound I screened today, so that it can bind to the enzyme. In the next days I will do the X-ray crystallography experiment to solve the structure of the complex.
Sometimes we use an in-house system like the one in the picture, a diffractometer that emits X-rays on the crystals and collects the diffraction patterns. Most of the time, this type of experiments is performed at the Oxford synchrotron, a particle accelerator where electrons are accelerated in a massive circular tunnel and used to shoot crystals. I know, I am getting very technical… maybe in my next post I will tell you more about this awesome technique!
I am done in the lab for today. I finally can go to the gym and relieve a bit of stress on the tread mill. Moreover, I get to properly train for the half marathon I am going to run in September with some of the Randall pals.
Another day is gone. Now dinner, catch up with my favourite series and sleep!
Written by: Giancarlo Abis