| AVS 63rd International Symposium & Exhibition | |
| Nanometer-scale Science and Technology | Thursday Sessions |
| Session NS+BI-ThA |
| Session: | Applied Nanoscale Microscopy Techniques/Biomaterial Interfaces – New Advances |
| Presenter: | John Stephen Fletcher, University of Gothenburg, Sweden |
| Authors: | J.S. Fletcher, University of Gothenburg, Sweden P. Wehrli, University of Gothenburg, Sweden A. Farewell, University of Gothenburg, Seweden T.B. Angerer, University of Gothenburg, Sweden J. Gottfries, University of Gothenburg, Sweden |
| Correspondent: | Click to Email |
For many years ToF-SIMS has shown the promise of delivering new information of direct relevance to biological research. However, inadequacies in the ability to generate intact molecular ion species and then detect them with precise mass resolution and accuracy have held the technique back. Recent advances in ToF-SIMS, through the implementation of gas cluster ion beams (GCIBs) coupled to non-conventional MS systems, now permit the analysis of higher mass species from native, underivatised, biological specimen i.e. intact bacterial cells. Being able to characterise and understand changes in bacterial biochemistry as a result of environmental, biological or pharmacological stress is critical to address the global challenge of antibiotic resistance. For example, E. coli is able to rapidly adjust the biophysical properties of its membrane phospholipids to adapt to environmental challenges including starvation stress. Here, these membrane lipid modifications were investigated in glucose starved E. coli cultures and compared to a DrelADspoT (ppGpp0) mutant strain of E. coli, deficient in the stringent response, by means of time-of-flight secondary ion mass spectrometry (ToF-SIMS Cultures in stationary phase were found to exhibit a radically different lipid composition as compared to cultures in exponential growth phase. Wild-type E. coli reacted upon carbon starvation by lipid modifications including elongation, cyclopropanation and increased cardiolipin formation. Observations are consistent with variants of cardiolipins (CL), phosphatidylglycerols (PG), phosphatidylethanolamines (PE), phosphatidic acids (PA), and fatty acids. Notably, despite having a proteomic profile and a gene expression profile somewhat similar to the wild-type during growth, the ppGpp0 mutant E. coli strain was found to exhibit modified phospholipids corresponding to unsaturated analogues of those found in the wild-type. We concluded that the ppGpp0 mutant reacts upon starvation stress by elongation and desaturation of fatty acyl chains, implying that only the last step of the lipid modification, the cyclopropanation, is under stringent control. These observations suggest alternative stress response mechanisms and illustrate the role of the RelA and SpoT enzymes in the biosynthetic pathway underlying these lipid modifications.