Pioneering new technique may help scientists combat drug resistance in E.coli

Novel ‘crystal toothpaste’ technique draws on cutting-edge technology to study anti-microbial resistance

Crystals of E.coli membrane protein shine inside the syringe
Crystals of E.coli membrane protein shine inside the syringe
 
An international team of scientists are using a state-of-the-art technique that draws on the power of ‘big science’ machines to help investigate the problem of antimicrobial resistance in E.coli.
 
The team from Diamond Light Source, Imperial and McGill University in Canada, are investigating the structure of membrane proteins from E.coli cells. Membrane proteins are a particular important field for modern drug discovery. Cells contain an outer membrane that acts like a protective wall, shielding them from the outside, and proteins in the membrane carry things like nutrients and information from outside of the cell into it; this makes them very important targets for new drugs. Approximately 40% of drugs currently target proteins in the cell membrane, and they are of pivotal interest to scientists looking to design new antimicrobials.
 
Isabel De Moraes, who runs the Membrane Protein Laboratory at Diamond, is leading the investigation into the E.coli membrane proteins. She comments: “At the moment, we have a major problem in bacterial resistance. It’s becoming more and more difficulty to control infections. But if we know more about how these agents work then we’re in a better position to create antimicrobials capable of manipulating them and preventing infection.”
Isabel De Moraes testing the crystals at LCLS
Isabel De Moraes testing the crystals at LCLS
This international collaboration is studying membrane proteins involved in the process of energy transfer inside E.coli cells. The proteins are responsible for the energy flow through the membrane into the bacteria, allowing it to infect its host. Isabel comments: “We want to shut down the bacterial system. If we understand how the energy system works, we can control it. And without energy, the bacteria dies.”
 
To uncover this structure, the team have used a particularly novel approach. The technique involves crystallising proteins of the bacteria inside a syringe and then squeezing them out, like a tube of toothpaste. The ‘crystal toothpaste’ will be squeezed out into the path of an XFEL laser beam, which will travel through the E.coli protein crystals and produce detailed data on their structure. This ‘crystal toothpaste’ technique allows scientists to study more crystal samples than ever before, providing more data on the nature and function of E.coli.
 
The novel technique involves growing crystals inside a syringe: a feat that is even more complex than it sounds. The crystals were developed at Diamond over a period of four years. During this time, the team refined the proteins using synchrotron light on Diamond’s I24 beamline. This allowed them to test crystal after crystal, each time adjusting the chemical composition until they created the perfect crystal sample.
 
Danny Axford on the beamline at LCLS where the crystals were tested
Danny Axford on the beamline at LCLS where the crystals were tested
The tiny, delicate crystals of E.coli membrane protein are too fragile to be tested on Diamond’s beamlines – exposure to radiation will destroy them almost instantly, making it impossible to get enough data. So the crystals will be taken to the SLAC National Accelerator Laboratory at Stanford University. The laboratory houses the Linac Coherent Light Source (LCLS), the world’s most powerful X-ray laser, which produces extremely powerful and short-lived pulses of light. These very brief pulses last, not microseconds, but femtoseconds; in this miniscule amount of time, the team can gather vast amounts of data on the structure of the crystals before they can be destroyed.
 
Whilst still in its early stages, this project demonstrates the power of drawing on different forms of technology to create synergies within scientific research. Together, the LCLS in Stanford and Diamond Light Source are allowing scientists to trial exciting new techniques and explore cutting-edge approaches to pressing issues such as drug resistance.