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An LC-MS/MS assay and complementary web-based tool to quantify and predict compound accumulation in E. coli.

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Discovering new antibiotics is hard due to Gram-negative bacteria

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Area of Science:

  • Microbiology
  • Chemical Biology
  • Drug Discovery

Background:

  • Gram-negative bacterial membranes pose a significant barrier to antibiotic entry.
  • Understanding compound physicochemical properties is crucial for intracellular accumulation.
  • Existing methods for assessing accumulation have limitations.

Purpose of the Study:

  • To develop a robust method for assessing small molecule intracellular accumulation in Gram-negative bacteria.
  • To identify key physicochemical properties governing compound accumulation in E. coli.
  • To create a predictive tool for antibiotic accumulation.

Main Methods:

  • Utilized liquid chromatography-tandem mass spectrometry (LC-MS/MS) for label-free compound accumulation assays.
  • Developed the eNTRy rules based on unbiased accumulation trend analysis in Escherichia coli.
  • Created the eNTRyway web tool for calculating properties and predicting accumulation.

Main Results:

  • Established the eNTRy rules: compounds with an ionizable Nitrogen, low Three-dimensionality, and rigidity are likely to accumulate in E. coli.
  • Demonstrated the utility of LC-MS/MS assays for broad small molecule analysis.
  • Developed a user-friendly web tool (eNTRyway) for predicting compound accumulation.

Conclusions:

  • The eNTRy rules provide a framework for designing antibiotics that can penetrate Gram-negative bacteria.
  • The LC-MS/MS assay and eNTRyway tool offer a practical workflow for drug discovery laboratories.
  • This approach facilitates the discovery of novel broad-spectrum antibiotics.