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Rapid Identification of Secondary Structure and Binding Site Residues in an Intrinsically Disordered Protein Segment
Mycobacterium tuberculosis harbours nine toxin-antitoxin (TA) systems of the MazEF family. MazEF TA modules are of immense importance due to the perceived role of the MazF toxin in M. tuberculosis persistence and disease. The MazE antitoxin has a disordered C-terminal domain that binds the toxin, Ma...
Autores principales: | , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Frontiers Media S.A.
2021
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8593223/ https://www.ncbi.nlm.nih.gov/pubmed/34795695 http://dx.doi.org/10.3389/fgene.2021.755292 |
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author | Chandra, Soumyanetra Chattopadhyay, Gopinath Varadarajan, Raghavan |
author_facet | Chandra, Soumyanetra Chattopadhyay, Gopinath Varadarajan, Raghavan |
author_sort | Chandra, Soumyanetra |
collection | PubMed |
description | Mycobacterium tuberculosis harbours nine toxin-antitoxin (TA) systems of the MazEF family. MazEF TA modules are of immense importance due to the perceived role of the MazF toxin in M. tuberculosis persistence and disease. The MazE antitoxin has a disordered C-terminal domain that binds the toxin, MazF and neutralizes its endoribonuclease activity. However, the structure of most MazEF TA complexes remains unsolved till date, obscuring structural and functional information about the antitoxins. We present a facile method to identify toxin binding residues on the disordered antitoxin. Charged residue scanning mutagenesis was used to screen a yeast surface displayed MazE6 antitoxin library against its purified cognate partner, the MazF6 toxin. Binding residues were deciphered by probing the relative reduction in binding to the ligand by flow cytometry. We have used this to identify putative antitoxin interface residues and local structure attained by the antitoxin upon interaction in the MazEF6 TA system and the same methodology is readily applicable to other intrinsically disordered protein regions. |
format | Online Article Text |
id | pubmed-8593223 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-85932232021-11-17 Rapid Identification of Secondary Structure and Binding Site Residues in an Intrinsically Disordered Protein Segment Chandra, Soumyanetra Chattopadhyay, Gopinath Varadarajan, Raghavan Front Genet Genetics Mycobacterium tuberculosis harbours nine toxin-antitoxin (TA) systems of the MazEF family. MazEF TA modules are of immense importance due to the perceived role of the MazF toxin in M. tuberculosis persistence and disease. The MazE antitoxin has a disordered C-terminal domain that binds the toxin, MazF and neutralizes its endoribonuclease activity. However, the structure of most MazEF TA complexes remains unsolved till date, obscuring structural and functional information about the antitoxins. We present a facile method to identify toxin binding residues on the disordered antitoxin. Charged residue scanning mutagenesis was used to screen a yeast surface displayed MazE6 antitoxin library against its purified cognate partner, the MazF6 toxin. Binding residues were deciphered by probing the relative reduction in binding to the ligand by flow cytometry. We have used this to identify putative antitoxin interface residues and local structure attained by the antitoxin upon interaction in the MazEF6 TA system and the same methodology is readily applicable to other intrinsically disordered protein regions. Frontiers Media S.A. 2021-11-02 /pmc/articles/PMC8593223/ /pubmed/34795695 http://dx.doi.org/10.3389/fgene.2021.755292 Text en Copyright © 2021 Chandra, Chattopadhyay and Varadarajan. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Genetics Chandra, Soumyanetra Chattopadhyay, Gopinath Varadarajan, Raghavan Rapid Identification of Secondary Structure and Binding Site Residues in an Intrinsically Disordered Protein Segment |
title | Rapid Identification of Secondary Structure and Binding Site Residues in an Intrinsically Disordered Protein Segment |
title_full | Rapid Identification of Secondary Structure and Binding Site Residues in an Intrinsically Disordered Protein Segment |
title_fullStr | Rapid Identification of Secondary Structure and Binding Site Residues in an Intrinsically Disordered Protein Segment |
title_full_unstemmed | Rapid Identification of Secondary Structure and Binding Site Residues in an Intrinsically Disordered Protein Segment |
title_short | Rapid Identification of Secondary Structure and Binding Site Residues in an Intrinsically Disordered Protein Segment |
title_sort | rapid identification of secondary structure and binding site residues in an intrinsically disordered protein segment |
topic | Genetics |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8593223/ https://www.ncbi.nlm.nih.gov/pubmed/34795695 http://dx.doi.org/10.3389/fgene.2021.755292 |
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