Functional and Biochemical Characterization of the MazEF6 Toxin-Antitoxin System of Mycobacterium tuberculosis

Chattopadhyay, G and Bhasin, M and Ahmed, S and Priya Gosain, T and Ganesan, S and Das, S and Thakur, C and Chandra, N and Singh, R and Varadarajan, R (2022) Functional and Biochemical Characterization of the MazEF6 Toxin-Antitoxin System of Mycobacterium tuberculosis. In: Journal of Bacteriology, 204 (4).

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Abstract

The Mycobacterium tuberculosis genome harbors nine toxin-antitoxin (TA) systems that are members of the mazEF family, unlike other prokaryotes, which have only one or two. Although the overall tertiary folds of MazF toxins are predicted to be similar, it is unclear how they recognize structurally different RNAs and antitoxins with divergent sequence specificity. Here, we have expressed and purified the individual components and complex of the MazEF6 TA system from M. tuberculosis. Size exclusion chromatography- multiangle light scattering (SEC-MALS) was performed to determine the oligomerization status of the toxin, antitoxin, and the complex in different stoichiometric ratios. The relative stabilities of the proteins were determined by nano-differential scanning fluorimetry (nano-DSF). Microscale thermophoresis (MST) and yeast surface display (YSD) were performed to measure the relative affinities between the cognate toxin- antitoxin partners. The interaction between MazEF6 complexes and cognate promoter DNA was also studied using MST. Analysis of paired-end RNA sequencing data revealed that the overexpression of MazF6 resulted in differential expression of 323 transcripts in M. tuberculosis. Network analysis was performed to identify the nodes from the topresponse network. The analysis of mRNA protection ratios resulted in identification of putative MazF6 cleavage site in its native host, M. tuberculosis. © 2022 American Society for Microbiology. All rights reserved.

Item Type:	Journal Article
Publication:	Journal of Bacteriology
Publisher:	American Society for Microbiology
Additional Information:	The copyright for this article belongs to American Society for Microbiology
Keywords:	antitoxin; bacterial protein, genetics; human; metabolism; Mycobacterium tuberculosis; toxin-antitoxin system; tuberculosis, Antitoxins; Bacterial Proteins; Humans; Mycobacterium tuberculosis; Toxin-Antitoxin Systems; Tuberculosis
Department/Centre:	Division of Biological Sciences > Biochemistry Division of Biological Sciences > Molecular Biophysics Unit
Date Deposited:	20 May 2022 09:00
Last Modified:	20 May 2022 09:00
URI:	https://eprints.iisc.ac.in/id/eprint/72044

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