Related Experiment Video
Updated: May 7, 2026
Preparation of Mycobacterium tuberculosis Culture Filtrate to Understand TB Pathogenesis
Published on: March 28, 2025
Proteomic analysis of Mycobacterium tuberculosis isolates resistant to kanamycin and amikacin
Bhavnesh Kumar1, Divakar Sharma, Prashant Sharma
1Department of Biochemistry, National JALMA Institute for Leprosy & Other Mycobacterial Diseases (Indian Council of Medical Research), Dr. Matsuki Miyazaki Road, Tajganj, PO Box No-1101, Agra, PIN-282004 India.
Abstract:
Kanamycin (KM) and amikacin (AK) are the key aminoglycoside drugs against tuberculosis (TB) and resistance to them severely affects the options for treatment. Many explanations have been proposed for drug resistance to these drugs but still some mechanisms are unknown. Proteins are the functional moiety of the cell and manifest in most of the biological processes; so, these are potential foci for the development of new therapeutics, diagnostics and vaccine. We examined the KM and AK resistant isolates of Mycobacterium tuberculosis using proteomic analysis comprising of two dimensional gel electrophoresis (2DGE), matrix assisted laser desorption ionization time-of-flight/time-of flight (MALDI-TOF/TOF) and bioinformatic tools like BLASTP, InterProScan, KEGG motif scan and molecular docking. Proteins intensities of twelve spots were found to be consistently increased in KM and AK resistant isolates and these were identified as Rv3867, Rv1932, Rv3418c, Rv1876, Rv2031c, Rv0155, Rv0643c, Rv3224, Rv0952, and Rv0440. Among these, Rv3867 and Rv3224 were identified as proteins with unknown function. All the proteins identified were cellular proteins. Molecular docking shows the proper interaction of both drugs with these molecules. Also, Rv1876 and Rv3224 were found to be probably involved in iron regulation/metabolism indicating the role of iron in imparting resistance to second line drugs.
Biological Significance:
The study that was carried out shows that two dimensional electrophoresis along with mass spectrometry is still the best approach for proteomic analysis. To the best of our knowledge it is the first ever report on proteomic analysis of M. tuberculosis isolates resistant to second line drugs (kanamycin and amikacin). The major finding implicates that the genes/proteins involved in iron metabolism and the two hypothetical proteins (Rv3867 and Rv3224) might be playing some crucial role in contributing resistance to second line drugs. Further exploitation in this direction may lead to the development of newer therapeutics against tuberculosis.
Insights
Drug resistance in tuberculosis (TB) is a growing concern. This study used proteomics to identify proteins, including those involved in iron metabolism and two hypothetical proteins, that may contribute to kanamycin and amikacin resistance in Mycobacterium tuberculosis.
Area of Science:
- Microbiology
- Proteomics
- Drug Resistance
Background:
- Kanamycin and amikacin are crucial aminoglycoside drugs for tuberculosis (TB) treatment.
- Resistance to these drugs severely limits therapeutic options.
- Understanding resistance mechanisms is vital for developing new treatments.
Purpose of the Study:
- To investigate the proteomic changes associated with kanamycin and amikacin resistance in Mycobacterium tuberculosis isolates.
- To identify potential protein targets involved in aminoglycoside resistance.
Main Methods:
- Proteomic analysis using two-dimensional gel electrophoresis (2DGE) and MALDI-TOF/TOF mass spectrometry.
- Bioinformatic tools including BLASTP, InterProScan, and KEGG motif scan.
- Molecular docking to assess drug-protein interactions.
Main Results:
- Twelve protein spots showed increased intensity in resistant isolates, including Rv3867, Rv1932, Rv3418c, Rv1876, Rv2031c, Rv0155, Rv0643c, Rv3224, Rv0952, and Rv0440.
- Rv3867 and Rv3224 were identified as proteins with unknown functions.
- Rv1876 and Rv3224 were implicated in iron regulation/metabolism, suggesting a role for iron in resistance.
Conclusions:
- Two-dimensional electrophoresis and mass spectrometry remain effective for proteomic analysis.
- This study is the first to report proteomic analysis of Mycobacterium tuberculosis isolates resistant to second-line drugs (kanamycin and amikacin).
- Proteins involved in iron metabolism and two hypothetical proteins (Rv3867, Rv3224) may play a significant role in conferring resistance to these drugs, offering potential targets for new TB therapeutics.
