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Dynamic Measurement of Protein Translation in Mycobacteria Using Nontargeted Stable Isotope Labeling in Combination
Christoph Haisch1, Anna-Cathrine Neumann-Cip2,3,4, Axel Imhof5
1Chair of Analytical Chemistry, Technical University of Munich, 85748 Garching, Germany.
Analytical Chemistry
|February 25, 2025
Summary
This study introduces a new proteomics method to dynamically measure protein synthesis in mycobacteria. This approach aids in discovering new tuberculosis drugs and predicting synergistic antibiotic combinations.
Area of Science:
- Microbiology
- Proteomics
- Drug Discovery
Background:
- Tuberculosis (TB) remains a major global health threat, necessitating new therapeutic strategies.
- Understanding mycobacterial metabolic pathways is crucial for developing novel antibiotics and effective TB treatment regimens.
Purpose of the Study:
- To present a novel proteomics approach for dynamically measuring nascent protein fractions in mycobacteria.
- To assess the impact of various antimicrobials on de novo protein synthesis.
Main Methods:
- Utilized nontargeted stable isotope incorporation (glycerol-1,3-13C2) to label nascent proteins.
- Employed matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF-MS) with a custom script for quantitative analysis.
- Compared protein synthesis under normal conditions and in response to rifampicin, linezolid, bedaquiline, and benzithiazinone-043.
Main Results:
- Enabled high-resolution monitoring of protein synthesis changes within half a doubling cycle.
- Demonstrated the ability to track the onset and extent of antimycobacterial activity.
- Revealed the potential to predict synergistic effects of drug combinations.
Conclusions:
- The dynamic protein synthesis analysis serves as a rapid screening tool for antimycobacterial activity and regulatory phenotypes.
- This untargeted, non-genetic modification approach is applicable to both model strains and clinical isolates.
- The method offers insights into drug action kinetics and aids in the development of effective TB therapies.
