Click-based determination of accumulation of molecules in Escherichia coli.

ABSTRACT: Gram-negative bacterial pathogens pose a significant challenge in drug development because their outer membranes hinder the permeation of small molecules. The lack of widely adoptable methods for measuring the cytosolic accumulation of compounds in bacterial cells further hinders drug discovery efforts. To address this challenge, we report the development of the Chloroalkane Azide Membrane Permeability (CHAMP) assay, which we designed specifically to assess molecule accumulation in the cytosol of Gram-negative bacteria. The CHAMP analysis utilizes bioorthogonal epitopes anchored within HaloTag-expressing bacteria and measures the cytosolic arrival of azide-bearing test molecules through strain-promoted azide-alkyne cycloaddition. This workflow enables robust and rapid accumulation measurem

SUBMITTER: Ongwae GM 

PROVIDER: S-EPMC12936067 | biostudies-literature | 2026 Jan

REPOSITORIES: biostudies-literature