A TCD researcher makes the first use of X-rays to capture structural images of proteins that are essential for health, writes Dick Ahlstrom
If you want to study something very small you need a very bright light to see it. A research group at Trinity College Dublin used one of the brightest available, a powerful beam of X-rays, to discover the shape of a protein.
Dr Amir Khan in Trinity's school of biochemistry and immunology joined with colleagues at University College Cork to determine the shape of a particularly important cell protein with the cumbersome name, Rab-11-FIP2 complex.
Shape is important when it comes to proteins because shape provides information about function, he explains. "If you don't know what a car looks like inside, how can you know how it works?"
The Rab-FIP complex is two proteins entwined together, explains Dr Khan, who received a Science Foundation Ireland investigator grant to pursue this research. "They regulate cargo deliveries in cells. They move cargo around within the cells, things like receptors."
Receptors are dotted around the outside of cells and serve as docking sites for proteins and other substances. If something connects to the receptor this initiates some action by the cell.
The Rab-FIP complex is a particular target for researchers, says Dr Khan. "There are many molecules on the surface of the cell membrane. We are interested in this because there are links between Rab-11-FIP2 and cancer."
Disease can arise if the complex is misshapen due to mutations in the cellular machinery that produces it and if mutated it causes "anarchy within the cell", hence their interest in its shape.
When a biologist wants to determine the shape of a protein he turns to X-ray crystallography. This involves firing a stream of X-rays at a protein that has been crystallised and watching how the X-ray beam is diffracted.
"You can imagine sunlight lighting a curtain. It produces a diffracted pattern of light that allows you to visualise the flower patterns in the curtain," he explains.
Cell biologists at UCC studied what the protein does in the cell and got its DNA sequence, which Dr Khan in turned used to produce pure Rab-FIP protein. "We optimised the protein crystals. We can't do this with the proteins in a liquid state, it has to be a crystalline state."
Trinity was the first to install its own X-ray crystallography equipment and now the University of Limerick also has a system, he says.
The problem is the X-ray beam this equipment produces isn't powerful enough to produce the very clear diffraction patterns needed to study the Rab-FIP complex. The researchers therefore turned to the European Synchrotron Radiation Facility in Grenoble, France to get a stronger X-ray beam.
"Because the proteins are so small we need a very intense light to see them," Dr Khan explains. "The higher the intensity of the X-rays the more we can visualise the protein. We have a certain intensity here but there is a huge intensity in Grenoble. It is just like having a brighter light, a much more intense light."
Ireland is the only European state that is not a member of the ESRF, but Dr Khan gained access to the facility via assistance from another European research lab. He brought his crystallised Rab11-FIP2 to Grenoble in December 2005.
Synchrotrons were built to allow physicists to smash atoms in the search for smaller and smaller fundamental particles. The ring-shaped chambers use powerful magnetic fields to accelerate electrons or positrons to close to the speed of light and guide them around the ring. They give off powerful X-rays in the process which can be directed at crystals. In this way the team became the first to identify the structure of the Rab11-FIP2 complex and they published their findings yesterday in the Cell Press journal, Structure.
With the precise structure of the complex now available, Dr Khan can use a new computer-driven imaging system at Trinity to produce large three-dimensional images of the protein's shape.
Installed with the financial support of the Higher Education Authority's Programme for Research in Third Level Institutions programme, the visualisation system uses rear projection to cast an image of the protein onto a screen. The researchers use special glasses, not unlike the old 3-D specs from the cinema, to produce an image that they can actually look into.
"It creates this false image of the structure that actually seems to come out of the screen," says Dr Khan.