21 October 2015
Jeremy Lakey and Tim Robinson (Newcastle University) use INTER to study model bacterial outer membranes in an earlier stage of the work
Scientists have a brand new tool for developing new antibiotics and other drugs in the fight against infections such as E.coli (Escherichia coli). 2013 figures taken from the NHS website show one in five infections involving E.coli is now resistant to a commonly-used antibiotic (ciproflaxin). Now researchers from Newcastle University and STFC have produced a working model of the outer membrane of E.coli, giving access to unprecedented information about the bacteria which is notoriously hard to study due to its size and exterior.
The results, published in the leading chemistry journal Angewandte Chemie International Edition, are featured as a ‘Research Highlight’ in Nature Chemical Biology. (20 October 2015).
It is well known that all types of disease-causing bacteria are becoming resistant to antibiotics. Antimicrobial resistance has been named as the key challenge in the Longitude Prize, an initiative launched by the Prime Minister at G8 2014 with a £10 million prize fund to help solve one of the greatest issues of our time.
Single-celled Gram-negatives, the group of bacteria that E.coli belongs to, are of special concern because they have an extra wall around their cells which can protect them physically from our treatments.
“Our model of the bacterial outer membrane can be used as a simulator to test how antibiotic molecules can be made to cross this critical barrier”, Jeremy Lakey, Professor of Structural Biochemistry at Newcastle University who led the study explains.
“A stable model is so important because the detailed structure of this wall is still not clear, largely because bacteria are very small and have a protective envelope that is only 20 nanometres thick. This model gives us unprecedented access to the structure and dynamics of the membrane.”
Gram-negative bacteria are highly successful organisms. In evolutionary terms, they are believed to have descended from a common ancestor of cyanobacteria, which emerged 3.6 billion years ago. E. coli bacteria live in the digestive tract of people and animals and most are harmless. However, some Gram-negatives cause illnesses such as meningitis, plague, Legionnaires disease, cholera and food poisoning. Understanding the outer membranes of Gram-negative bacteria is important for antibiotic development but their structure and dynamics are poorly understood because of their small size and inaccurate lab, or in vitro, models.
The nanoscale structure of the membrane was determined using STFC’s ISIS Neutron and Muon source. An instrument called POLREF allowed the precise molecular composition to be resolved, showing accurate details of the model such as the molecular asymmetry and the thickness of its internal water layer.
“We now are able to undertake studies on model bacterial membranes under conditions which much more closely resemble those found of live bacteria than has previous been possible”, said Luke Clifton from ISIS. “Neutron scattering allows us to resolve complex structures composed of mixtures of biomolecules. By combining this with isotopic labelling, to which neutrons are very sensitive, we were able to determine where each component of the model was”.
Clifton and co-workers went on to test the response of the model to antimicrobial proteins produced in our bodies, including lysozyme and lactoferrin. Interactions of these proteins with the outer membrane in vivo and in vitro are well known, allowing for direct comparisons with the synthetic model. Neutron reflectivity revealed that the experiments reproduced in vivo behaviour, replicating the disruption of the outer membrane previously seen in living bacteria.
Jeremy Lakey said: “Biological cells are hugely complicated and we are continually trying new ways to understand their molecular structure. Neutrons are now an essential tool in this effort because they have the unique ability to both describe structure and tell the difference between the different types of molecules found in cells. Thus, in our studies of the bacterial envelope, moving to neutrons was like switching from black and white to colour TV.”
The next challenge for the researchers is to begin incorporating membrane proteins into the bilayer.
This work was funded by the Wellcome Trust.
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The ISIS pulsed neutron and muon source at STFC’s Rutherford Appleton Laboratory in Oxfordshire is a world-leading centre for research in the physical and life sciences. It is owned and operated by the Science and Technology Facilities Council.
ISIS produces beams of neutrons and muons that allow scientists to study materials at the atomic level using a suite of instruments, often described as ‘super-microscopes’. It supports a national and international community of more than 2000 scientists who use neutrons and muons for research in physics, chemistry, materials science, geology, engineering and biology. It is the most productive research centre of its type in the world.
From the original vision over 30 years ago, ISIS has become one of the UK’s major scientific achievements. As the world’s leading pulsed neutron and muon source, ISIS has changed the way the world views neutron scattering.
Polref is a polarised neutron reflectometer designed for the study of the magnetic ordering in and between the layers and surfaces of thin film materials.