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Chemistry for Prosperity: Green fluorescent protein in medicine

by Royal Society of Chemistry

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One of a series of case studies compiled by the Royal Society of Chemistry, this presentation tells the story of Green Fluorescent Protein and its uses in medicine. Please feel free to use this ...

One of a series of case studies compiled by the Royal Society of Chemistry, this presentation tells the story of Green Fluorescent Protein and its uses in medicine. Please feel free to use this information and help us to make the case for investing in the chemical sciences. Download the PowerPoint file to access the associated script.

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[Speaker introduces him/herself ] In September 2010 the RSC published a report showing that the chemistry research enables us to generate £258bn each year, or 21% of the UK’s GDP. An impressive figure but you may well wonder how this works? This case study, which is one of a series, attempts to answer that question by looking at the challenges that chemical scientists typically face, the solutions they have found and the wealth that this has created.
Cats that glow in the dark. Science fiction? Actually....no. These cats have each had a completely harmless fluorescent protein introduced into their genes. Why would someone do that? Well, this type of experiment is known as a proof-of-concept, and it has led to the development of an incredibly versatile tool in medicine.
Green fluorescent protein, or GFP, was first isolated from the jellyfish, Aequorea victoria, in the 1960s. Over the next decade, Japanese organic chemist and marine biologist, Osamu Shimomura (who took this photograph), studied its properties. However, it wasn’t until 1992, when American molecular biologist Douglas Prasher identified the genetic code of GFP and managed to clone it (or copy it), that scientists began to realize its potential for molecular biology. Two other research groups, led by Martin Chalfie and Frederick Tsuji, took things a step further by putting the GFP gene into the bacterium in E. coli and in the roundworm C. elegans, both of which produced the GFP protein. When an organism produces a protein this is in the form of a long chain of amino acids which must be folded into a specific shape for the protein to be active. Remarkably, the GFP molecule produced by the bacterium and the roundworm folded correctly and was fluorescent at room temperature. However, there were drawbacks including sensitivity to different levels of acidity, poor fluorescence, instability to light and poor folding at body temperatures (37°C). Over the following years, a number of mutants were developed including enhanced GFP (EGFP), which allowed the practical use of GFPs in the cells of mammals cells and “superfolder GFP”, which had a number of genetic mutations that allow GFP to rapidly fold and become flourescent, even under difficult conditions.
One of the areas in which GFP has made real impact is in understanding biological interactions associated with infection. All cells continuously display the proteins they have inside them on their surfaces. This is done by a group of molecules called the 'Major Histo-compatibility Complex’ or MHC. The immune system’s T-cells check the proteins on the surface of the cell for any which should not be there, identified here as a 'foreign object‘. The T-cell recognises that it should not be there, identifying this as an infected cell, which the immune system then destroys. Some viruses have developed the ability to defeat this process by removing the MHC, so that no proteins are displayed on the cell’s surface, so the T-cell does not recognise the cell as infected. Luckily, mammals have evolved a second line of defence, the 'Natural Killer Cell', which checks for the presence of MHC, not the proteins. If the cell surface is found not to contain MHC, the natural killer cell destroys the cell. By labelling the MHC with GFP, Dan Davis’ research group at UCL and others have been able to probe the fine details of the interaction of the MHC and the Natural Killer Cell. Understanding molecular recognition by Natural Killer Cells is of key importance for improving our understanding of the causes, development and effects, or the pathology, of viruses such as HIV.
GFP, among other proteins, is used in imaging techniques such as dual colour localisation microscopy, shown here imaging the detail of a bone cancer cell. The image on the left shows normal fluorescence microscopy, while that on the right uses the dual colour technique, differentiating between over 120,000 molecules. A high density of visible molecules is important in recognizing, for example, molecular clusters as sites of increased activity – an indication that there may be a problem. Currently, this technique is enabling major advances in pharmacology, cardiology and stem cell research. This is an excellent example of ground-breaking research leading to the development of tools which improve our understanding of disease and, ultimately, lead to new treatments.
Another fascinating use of GFP is termed Brainbow - a process by which individual neurons in the brain can be distinguished from neighbouring neurons using fluorescent proteins. Different ratios of red, green, and blue derivatives of green fluorescent protein are produced by individual neurons, giving each neuron a distinctive colour. This process has been a major contribution to the study of neural connections in the brain. The first demonstration of this technique was published in Nature in 2007 and was the work of Jeff W. Lichtman and Joshua R. Sanes at Harvard Medical School. Using Brainbow, researchers are now able to construct maps of neural circuits and investigate how these relate to various mental activities. Although most in vivo tests (in live organisms) to date have been done in mice, scientists are investigating using the technique in humans where it could help them to understand of the causes of neurological and psychological conditions. Green fluorescent protein was first isolated by a chemist who was driven by curiosity and for the next three decades this work had little impact outside of a research lab. Nowadays, GFP and numerous related proteins are used by biologists and medical specialists. A wide variety of colours can be produced in specific cells and scientists are only just beginning to understand the potential of this amazing tool.

Chemistry for Prosperity: Green fluorescent protein in medicine Presentation Transcript

Chemistry for ProsperityGreen fluorescent protein in medicine
© Osamu Shimomura
© Christoph Crem
Chemistry for ProsperityGreen fluorescent protein in medicine