Ethambutol targets the glutamate racemase of Mycobacterium tuberculosis—an enzyme involved in peptidoglycan biosynthesis
Increasing drug resistance in pathogens including Mycobacterium tuberculosis (MTB) has been ascribed to mutations in the known target genes. However, many of these drugs have multiple targets; some of which have not been identified so far. Understanding the mechanism of action of these drugs holds a great promise in better management of disease especially by drug-resistant strains. In this study, we report glutamate racemase (MurI), a crucial enzyme of phase I peptidoglycan (PG) biosynthesis pathway of MTB, as an additional target of ethambutol (EMB). The effect on EMB on the MurI protein at structural and functional level was studied using different spectroscopic, biochemical, and insilico approaches. Spectroscopic analysis revealed that EMB-modified protein undergoes conformational alterations. Furthermore, in vitro racemization studies of the MurI protein suggest that EMB decreases its functional activity. Docking studies revealed that EMB interacts with most of the active residues at the binding site and blocks the binding pocket. Overall, data suggests that EMB, a primary drug used for the treatment of tuberculosis (TB), acts as a competitive inhibitor of substrate for binding to mycobacterial MurI protein. The study also points out to our lacunae in understanding the site and mechanism of action of existing drugs. Furthermore, glutamate racemase is a conserved protein of the bacterial kingdom; therefore, ethambutol could be a promising candidate as a broad-spectrum antibiotic for many other bacterial diseases.
KeywordsMycobacterium tuberculosis Glutamate racemase MurI Peptidoglycan Ethambutol d-glutamate
The authors thank the Department of Science and Technology (DST) purse grant and MG grant to DS for financial support. The support from Department of Biotechnology, Govt. of India for Bioinformatics Facility at Dr. B.R. Ambedkar Center for Biomedical Research is highly acknowledged. Senior research fellowship to AP from Indian Council for Medical Research and DST-INSPIRE fellowship to PJ from DST is gratefully acknowledged.
The project grants, purse grant and MG grant, were received by DST and University of Delhi. Fellowship and contingency grant was provided to AP from ICMR and PJ from DST.
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Conflict of interest
The authors declare that they have no conflict of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
- Belanger AE, Besra GS, Ford ME, Mikusova K, Belisle JT, Brennan PJ, Inamine JM (1996) The embAB genes of Mycobacterium avium encode an arabinosyl transferase involved in cell wall arabinan biosynthesis that is the target for the antimycobacterial drug ethambutol. Proc Natl Acad Sci 93:11919–11924. https://doi.org/10.1073/pnas.93.21.11919 CrossRefGoogle Scholar
- Gagnon JK, Law SM, Brooks III CL (2014) Flexible CDOCKER: development and application of a pseudo-explicit structure-based docking method within CHARMM. Biophys J 106:646a . doi: https://doi.org/10.1016/j.bpj.2013.11.3576
- Pronk S, Páll S, Schulz R, Larsson P, Bjelkmar P, Apostolov R, Shirts MR, Smith JC, Kasson PM, van der Spoel D, Hess B, Lindahl E (2013) GROMACS 4.5: a high-throughput and highly parallel open source molecular simulation toolkit. Bioinformatics 29:845–854. https://doi.org/10.1093/bioinformatics/btt055 CrossRefGoogle Scholar
- Raviglione M, Sulis G (2016) Tuberculosis 2015: burden, challenges and strategy for control and elimination. Infect Dis Rep 8. https://doi.org/10.4081/idr.2016.6570
- Tanwar O, Deora GS, Tanwar L, Kumar G, Janardhan S, Alam M, null S, Akhter M (2014) Novel hydrazine derivatives as selective DPP-IV inhibitors: findings from virtual screening and validation through molecular dynamics simulations. J Mol Model 20:2118. https://doi.org/10.1007/s00894-014-2118-7 CrossRefGoogle Scholar
- Walsh CT (1989) Enzymes in the D-alanine branch of bacterial cell wall peptidoglycan assembly. J Biol Chem 264:2393–2396Google Scholar