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...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2021-11-01
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| Series: | Frontiers in Genetics |
| Subjects: | |
| Online Access: | https://www.frontiersin.org/articles/10.3389/fgene.2021.755292/full |
| _version_ | 1818892277382119424 |
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| author | Soumyanetra Chandra Gopinath Chattopadhyay Raghavan Varadarajan |
| author_facet | Soumyanetra Chandra Gopinath Chattopadhyay Raghavan Varadarajan |
| author_sort | Soumyanetra Chandra |
| collection | DOAJ |
| 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. |
| first_indexed | 2024-12-19T17:54:09Z |
| format | Article |
| id | doaj.art-23f5baaf1da24b5198878081ecafdf31 |
| institution | Directory Open Access Journal |
| issn | 1664-8021 |
| language | English |
| last_indexed | 2024-12-19T17:54:09Z |
| publishDate | 2021-11-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Genetics |
| spelling | doaj.art-23f5baaf1da24b5198878081ecafdf312022-12-21T20:11:51ZengFrontiers Media S.A.Frontiers in Genetics1664-80212021-11-011210.3389/fgene.2021.755292755292Rapid Identification of Secondary Structure and Binding Site Residues in an Intrinsically Disordered Protein SegmentSoumyanetra ChandraGopinath ChattopadhyayRaghavan VaradarajanMycobacterium 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.https://www.frontiersin.org/articles/10.3389/fgene.2021.755292/fullcharged mutational scanningaspartatefunctional residuesinterfaceprotein-protein interactionsprotein structure |
| spellingShingle | Soumyanetra Chandra Gopinath Chattopadhyay Raghavan Varadarajan Rapid Identification of Secondary Structure and Binding Site Residues in an Intrinsically Disordered Protein Segment Frontiers in Genetics charged mutational scanning aspartate functional residues interface protein-protein interactions protein structure |
| 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 | charged mutational scanning aspartate functional residues interface protein-protein interactions protein structure |
| url | https://www.frontiersin.org/articles/10.3389/fgene.2021.755292/full |
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