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A hyperpromiscuous antitoxin protein domain for the neutralization of diverse toxin domains

Kurata, Tatsuaki LU ; Saha, Chayan Kumar LU orcid ; Buttress, Jessica A ; Mets, Toomas ; Brodiazhenko, Tetiana ; Turnbull, Kathryn J ; Awoyomi, Ololade F ; Oliveira, Sofia Raquel Alves ; Jimmy, Steffi and Ernits, Karin LU , et al. (2022) In Proceedings of the National Academy of Sciences of the United States of America 119(6). p.1-12
Abstract

Toxin-antitoxin (TA) gene pairs are ubiquitous in microbial chromosomal genomes and plasmids as well as temperate bacteriophages. They act as regulatory switches, with the toxin limiting the growth of bacteria and archaea by compromising diverse essential cellular targets and the antitoxin counteracting the toxic effect. To uncover previously uncharted TA diversity across microbes and bacteriophages, we analyzed the conservation of genomic neighborhoods using our computational tool FlaGs (for flanking genes), which allows high-throughput detection of TA-like operons. Focusing on the widespread but poorly experimentally characterized antitoxin domain DUF4065, our in silico analyses indicated that DUF4065-containing proteins serve as... (More)

Toxin-antitoxin (TA) gene pairs are ubiquitous in microbial chromosomal genomes and plasmids as well as temperate bacteriophages. They act as regulatory switches, with the toxin limiting the growth of bacteria and archaea by compromising diverse essential cellular targets and the antitoxin counteracting the toxic effect. To uncover previously uncharted TA diversity across microbes and bacteriophages, we analyzed the conservation of genomic neighborhoods using our computational tool FlaGs (for flanking genes), which allows high-throughput detection of TA-like operons. Focusing on the widespread but poorly experimentally characterized antitoxin domain DUF4065, our in silico analyses indicated that DUF4065-containing proteins serve as broadly distributed antitoxin components in putative TA-like operons with dozens of different toxic domains with multiple different folds. Given the versatility of DUF4065, we have named the domain Panacea (and proteins containing the domain, PanA) after the Greek goddess of universal remedy. We have experimentally validated nine PanA-neutralized TA pairs. While the majority of validated PanA-neutralized toxins act as translation inhibitors or membrane disruptors, a putative nucleotide cyclase toxin from a Burkholderia prophage compromises transcription and translation as well as inducing RelA-dependent accumulation of the nucleotide alarmone (p)ppGpp. We find that Panacea-containing antitoxins form a complex with their diverse cognate toxins, characteristic of the direct neutralization mechanisms employed by Type II TA systems. Finally, through directed evolution, we have selected PanA variants that can neutralize noncognate TA toxins, thus experimentally demonstrating the evolutionary plasticity of this hyperpromiscuous antitoxin domain.

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organization
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type
Contribution to journal
publication status
published
subject
in
Proceedings of the National Academy of Sciences of the United States of America
volume
119
issue
6
article number
e2102212119
pages
1 - 12
publisher
National Academy of Sciences
external identifiers
  • scopus:85124146792
  • pmid:35121656
ISSN
1091-6490
DOI
10.1073/pnas.2102212119
language
English
LU publication?
yes
additional info
Copyright © 2022 the Author(s). Published by PNAS.
id
6186b035-c416-4952-b9f8-302fab461e4b
date added to LUP
2022-02-09 17:14:36
date last changed
2024-04-18 05:18:19
@article{6186b035-c416-4952-b9f8-302fab461e4b,
  abstract     = {{<p>Toxin-antitoxin (TA) gene pairs are ubiquitous in microbial chromosomal genomes and plasmids as well as temperate bacteriophages. They act as regulatory switches, with the toxin limiting the growth of bacteria and archaea by compromising diverse essential cellular targets and the antitoxin counteracting the toxic effect. To uncover previously uncharted TA diversity across microbes and bacteriophages, we analyzed the conservation of genomic neighborhoods using our computational tool FlaGs (for flanking genes), which allows high-throughput detection of TA-like operons. Focusing on the widespread but poorly experimentally characterized antitoxin domain DUF4065, our in silico analyses indicated that DUF4065-containing proteins serve as broadly distributed antitoxin components in putative TA-like operons with dozens of different toxic domains with multiple different folds. Given the versatility of DUF4065, we have named the domain Panacea (and proteins containing the domain, PanA) after the Greek goddess of universal remedy. We have experimentally validated nine PanA-neutralized TA pairs. While the majority of validated PanA-neutralized toxins act as translation inhibitors or membrane disruptors, a putative nucleotide cyclase toxin from a Burkholderia prophage compromises transcription and translation as well as inducing RelA-dependent accumulation of the nucleotide alarmone (p)ppGpp. We find that Panacea-containing antitoxins form a complex with their diverse cognate toxins, characteristic of the direct neutralization mechanisms employed by Type II TA systems. Finally, through directed evolution, we have selected PanA variants that can neutralize noncognate TA toxins, thus experimentally demonstrating the evolutionary plasticity of this hyperpromiscuous antitoxin domain.</p>}},
  author       = {{Kurata, Tatsuaki and Saha, Chayan Kumar and Buttress, Jessica A and Mets, Toomas and Brodiazhenko, Tetiana and Turnbull, Kathryn J and Awoyomi, Ololade F and Oliveira, Sofia Raquel Alves and Jimmy, Steffi and Ernits, Karin and Delannoy, Maxence and Persson, Karina and Tenson, Tanel and Strahl, Henrik and Hauryliuk, Vasili and Atkinson, Gemma C}},
  issn         = {{1091-6490}},
  language     = {{eng}},
  number       = {{6}},
  pages        = {{1--12}},
  publisher    = {{National Academy of Sciences}},
  series       = {{Proceedings of the National Academy of Sciences of the United States of America}},
  title        = {{A hyperpromiscuous antitoxin protein domain for the neutralization of diverse toxin domains}},
  url          = {{http://dx.doi.org/10.1073/pnas.2102212119}},
  doi          = {{10.1073/pnas.2102212119}},
  volume       = {{119}},
  year         = {{2022}},
}