Cell wall enrichment unveils proteomic changes in the cell wall during treatment of Mycobacterium smegmatis with sub-lethal concentrations of rifampicin

Journal of Proteomics
Clemens HermannJonathan M Blackburn

Abstract

Understanding the cell wall of mycobacteria is crucial for improving drug design or identifying new antigens suitable to vaccination. Yet this remains problematic due to the complexity of the cell wall composition. In this study, we successfully developed gel-free approaches to study cell wall proteins in Mycobacterium smegmatis. The cell wall was subjected to differential centrifugation, differential detergent solubilisation and phase separation to yield the genuine cell wall proteome. Next, protein extracts were digested by filter-assisted sample preparation for LC-MS/MS analysis on a Q Exactive mass spectrometer, and identified proteins filtered through a stringent bioinformatics pipeline. This yielded the unprecedented coverage of 96 lipoproteins, 475 membrane proteins and 73 secreted proteins. Employing this approach, we next quantified changes in the cell wall proteome during exposure of M. smegmatis to sub-lethal concentration of rifampicin. This facilitated detailed characterisation of the dysregulation of ABC transporters, virulence factors such as Mce proteins and PknG, and proteins involved in cell wall and lipid synthesis. Crucially, these cell wall proteins are under-represented in previous proteome analysis of M. ...Continue Reading

Citations

Dec 17, 2019·Molecular & Cellular Proteomics : MCP·Seanantha S BarosNelson C Soares
Jun 20, 2020·The Journal of Biological Chemistry·Laurie ThouvenelChristian Chalut
Jan 17, 2021·Molecular & Cellular Proteomics : MCP·Seanantha S BarosNelson C Soares
Jul 28, 2019·Microbiology Spectrum·Gabriel T MashabelaDigby F Warner
Nov 4, 2020·Journal of Proteome Research·Clemens HermannNelson C Soares

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