| Institution: | 1. Department of Chemistry, University of Virginia, 22904 Charlottesville, USA;2. Molecular and Cellular Biology Graduate Program, University of Massachusetts, Amherst, USA
Department of Microbiology, University of Massachusetts, Amherst, USA;3. Institute of Translational Medicine, Zhangjiang Institute for Advanced Study, Shanghai Jiao Tong University, 200232 Shanghai, China |
| Abstract: | The general lack of permeability of small molecules observed for Mycobacterium tuberculosis (Mtb) is most ascribed to its unique cell envelope. More specifically, the outer mycomembrane is hypothesized to be the principal determinant for access of antibiotics to their molecular targets. We describe a novel assay that combines metabolic tagging of the peptidoglycan, which sits directly beneath the mycomembrane, click chemistry of test molecules, and a fluorescent labeling chase step, to measure the permeation of small molecules. We showed that the assay workflow was robust and compatible with high-throughput analysis in mycobacteria by testing a small panel of azide-tagged molecules. The general trend is similar across the two types of mycobacteria with some notable exceptions. We anticipate that this assay platform will lay the foundation for medicinal chemistry efforts to understand and improve uptake of both existing drugs and newly-discovered compounds into mycobacteria. |
