Colistin kills bacteria by targeting lipopolysaccharide in the cytoplasmic membrane
- Imperial College London, United Kingdom
- University of Texas at Austin, United States
Abstract
Colistin is an antibiotic of last resort, but has poor efficacy and resistance is a growing problem. Whilst it is well established that colistin disrupts the bacterial outer membrane by selectively targeting lipopolysaccharide (LPS), it was unclear how this led to bacterial killing. We discovered that MCR-1 mediated colistin resistance in Escherichia coli is due to modified LPS at the cytoplasmic rather than outer membrane. In doing so, we also demonstrated that colistin exerts bactericidal activity by targeting LPS in the cytoplasmic membrane. We then exploited this information to devise a new therapeutic approach. Using the LPS transport inhibitor murepavadin, we were able to cause LPS accumulation in the cytoplasmic membrane of Pseudomonas aeruginosa, which resulted in increased susceptibility to colistin in vitro and improved treatment efficacy in vivo. These findings reveal new insight into the mechanism by which colistin kills bacteria, providing the foundations for novel approaches to enhance therapeutic outcomes.
Data availability
Source data for all figures has been deposited at Dryad: https://doi.org/10.5061/dryad.98sf7m0hh
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Data from: Colistin kills bacteria by targeting lipopolysaccharide in the cytoplasmic membraneDryad Digital Repository, doi: 10.5061/dryad.98sf7m0hh.
Article and author information
Author details
R Christopher D Furniss
Funding
Medical Research Council (PhD Studentship)
- Akshay Sabnis
Wellcome Trust
- Andrew M Edwards
NIHR Imperial Biomedical Research Centre
- Andrew M Edwards
DFG
- Anna Klöckner
Horizon 2020
- Anna Klöckner
Rosetrees Trust
- Molly M Stevens
Cystic Fibrosis Trust
- Jane C Davies
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: The use of mice was performed under the authority of the UK Home Office outlined in the Animals (Scientific Procedures) Act 1986 after ethical review by Imperial College London Animal Welfare and Ethical Review Body (PPL 70/7969).
Copyright
© 2021, Sabnis et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Colistin kills bacteria by targeting lipopolysaccharide in the cytoplasmic membrane
eLife 10:e65836.
https://doi.org/10.7554/eLife.65836
Further reading
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Glutamine synthetases (GS) are central enzymes essential for the nitrogen metabolism across all domains of life. Consequently, they have been extensively studied for more than half a century. Based on the ATP-dependent ammonium assimilation generating glutamine, GS expression and activity are strictly regulated in all organisms. In the methanogenic archaeon Methanosarcina mazei, it has been shown that the metabolite 2-oxoglutarate (2-OG) directly induces the GS activity. Besides, modulation of the activity by interaction with small proteins (GlnK1 and sP26) has been reported. Here, we show that the strong activation of M. mazei GS (GlnA1) by 2-OG is based on the 2-OG dependent dodecamer assembly of GlnA1 by using mass photometry (MP) and single particle cryo-electron microscopy (cryo-EM) analysis of purified strep-tagged GlnA1. The dodecamer assembly from dimers occurred without any detectable intermediate oligomeric state and was not affected in the presence of GlnK1. The 2.39 Å cryo-EM structure of the dodecameric complex in the presence of 12.5 mM 2-OG demonstrated that 2-OG is binding between two monomers. Thereby, 2-OG appears to induce the dodecameric assembly in a cooperative way. Furthermore, the active site is primed by an allosteric interaction cascade caused by 2-OG-binding towards an adaption of an open active state conformation. In the presence of additional glutamine, strong feedback inhibition of GS activity was observed. Since glutamine dependent disassembly of the dodecamer was excluded by MP, feedback inhibition most likely relies on the binding of glutamine to the catalytic site. Based on our findings, we propose that under nitrogen limitation the induction of M. mazei GS into a catalytically active dodecamer is not affected by GlnK1 and crucially depends on the presence of 2-OG.
