Electric charge against antimicrobial resistance

A group of scientists working on antimicrobial resistance, a global problem that has recently been announced as the challenge set for the Longitude Prize, have used Diamond’s Circular Dichroism beamline to further understand the mechanisms involved.

An article published using data obtained from the Circular Dichroism beamline, B23, at Diamond Light Source has provided insights into the methods by which an inner membrane transporter, SbmA, functions. The results, published as a cover article in the Journal of Bacteriology, show how peptides produced by the innate immune response, Bac7, can interact and be transported by the bacterial protein SmbA. Upon internalisation, these peptides can kill the bacterial cells. These findings will help scientists to understand how to improve antimicrobial drugs to penetrate bacterial cells efficiently.

The World Health Organization estimates that antibiotics treatments add an average of 20 years to all of our lives. But in the 80 years since the discovery of penicillin, our overuse of antibiotics has put pressure on bacteria to evolve resistance, leading to the emergence of untreatable superbugs that threaten the basis of modern medicine.Previously, several studies have shown that SbmA can help to deliver antibacterial peptides – broad-spectrum antibiotics – into gram-negative bacteria, which are more likely to be resistant to antibiotics. However, how this process was driven had not been fully understood until now.

Using data obtained on B23, the researchers, from Imperial College London and the University of Trieste, Italy, were able to measure the interactions between the Bac7 peptide and SmbA.

Effect of Bac7 on the secondary structure and interaction on SbmA with binding constant of 300nM

“For the bacterial transporter SbmA, the natural substrates are at present unknown,” explains Dr Marco Scocchi of the University of Trieste, “However the antimicrobial peptide Bac7 exploits this membrane protein to penetrate into the bacterial cell.”

The researchers have been able to show that this process is proton-driven, rather than ATP driven. This means that due to a difference in the number of protons, the variance in pH and electric charge creates an electrochemical potential. This works similarly to that of a battery, storing energy for the cell.

The researchers found the facilities at Diamond Light Source extremely valuable to their project. “Studies [such as this] can be very challenging, and in the past have been very protein ‘hungry’,” said Dr Konstantinos Beis, of Imperial College London. “By using B23 we only needed very small amounts of protein to achieve data of such high quality. The sensitivity and accurate measurements from B23 have greatly enhanced our work.”

By understanding the methods of transport for antibacterial peptides into gram-negative bacteria, and the affinity with which these bind to each other, the researchers hope to take some steps towards developing new drugs that bacteria have less resistance to.

“Thanks to B23 we showed that this peptide actually binds to SbmA and we could measure the affinity constant for this binding,” concludes Dr Marco Scocchi, “[Thus] reinforcing the data previously obtained via affinity chromatography and moving towards the development of new antimicrobial drugs.”

To find out more about using the B23 beamline, or to discuss potential applications, please contact Dr Rohanah Hussain: rohanah.hussain@diamond.ac.uk

Runti G., Carmen Lopez Ruiz M. d., Stoilova T., Hussain R., Jennions M., Choudhury H. G., Benincasa M., Gennaro R., Beis K. and Scocci M. Functional Characterization of SbmA, a Bacterial Inner Membrane Transporter Required for Importing the Antimicrobial Peptide Bac7(1-35). Journal of Bacteriology 195, 5343-5351 (2013).