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Novel Toxin-Antitoxin Module SlvT-SlvA Regulates Megaplasmid Stability and Incites Solvent Tolerance in Pseudomonas putida S12

Pseudomonas putida S12 is highly tolerant of organic solvents in saturating concentrations, rendering this microorganism suitable for the industrial production of various aromatic compounds. Previous studies revealed that P. putida S12 contains the single-copy 583-kbp megaplasmid pTTS12. pTTS12 carr...

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Autores principales: Kusumawardhani, Hadiastri, van Dijk, David, Hosseini, Rohola, de Winde, Johannes H.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Society for Microbiology 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7301842/
https://www.ncbi.nlm.nih.gov/pubmed/32358012
http://dx.doi.org/10.1128/AEM.00686-20
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author Kusumawardhani, Hadiastri
van Dijk, David
Hosseini, Rohola
de Winde, Johannes H.
author_facet Kusumawardhani, Hadiastri
van Dijk, David
Hosseini, Rohola
de Winde, Johannes H.
author_sort Kusumawardhani, Hadiastri
collection PubMed
description Pseudomonas putida S12 is highly tolerant of organic solvents in saturating concentrations, rendering this microorganism suitable for the industrial production of various aromatic compounds. Previous studies revealed that P. putida S12 contains the single-copy 583-kbp megaplasmid pTTS12. pTTS12 carries several important operons and gene clusters facilitating P. putida S12 survival and growth in the presence of toxic compounds or other environmental stresses. We wished to revisit and further scrutinize the role of pTTS12 in conferring solvent tolerance. To this end, we cured the megaplasmid from P. putida S12 and conclusively confirmed that the SrpABC efflux pump is the major determinant of solvent tolerance on the megaplasmid pTTS12. In addition, we identified a novel toxin-antitoxin module (proposed gene names slvT and slvA, respectively) encoded on pTTS12 which contributes to the solvent tolerance phenotype and is important for conferring stability to the megaplasmid. Chromosomal introduction of the srp operon in combination with the slvAT gene pair created a solvent tolerance phenotype in non-solvent-tolerant strains, such as P. putida KT2440, Escherichia coli TG1, and E. coli BL21(DE3). IMPORTANCE Sustainable alternatives for high-value chemicals can be achieved by using renewable feedstocks in bacterial biocatalysis. However, during the bioproduction of such chemicals and biopolymers, aromatic compounds that function as products, substrates, or intermediates in the production process may exert toxicity to microbial host cells and limit the production yield. Therefore, solvent tolerance is a highly preferable trait for microbial hosts in the biobased production of aromatic chemicals and biopolymers. In this study, we revisit the essential role of megaplasmid pTTS12 from solvent-tolerant Pseudomonas putida S12 for molecular adaptation to an organic solvent. In addition to the solvent extrusion pump (SrpABC), we identified a novel toxin-antitoxin module (SlvAT) which contributes to short-term tolerance in moderate solvent concentrations, as well as to the stability of pTTS12. These two gene clusters were successfully expressed in non-solvent-tolerant strains of P. putida and Escherichia coli strains to confer and enhance solvent tolerance.
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spelling pubmed-73018422020-07-01 Novel Toxin-Antitoxin Module SlvT-SlvA Regulates Megaplasmid Stability and Incites Solvent Tolerance in Pseudomonas putida S12 Kusumawardhani, Hadiastri van Dijk, David Hosseini, Rohola de Winde, Johannes H. Appl Environ Microbiol Physiology Pseudomonas putida S12 is highly tolerant of organic solvents in saturating concentrations, rendering this microorganism suitable for the industrial production of various aromatic compounds. Previous studies revealed that P. putida S12 contains the single-copy 583-kbp megaplasmid pTTS12. pTTS12 carries several important operons and gene clusters facilitating P. putida S12 survival and growth in the presence of toxic compounds or other environmental stresses. We wished to revisit and further scrutinize the role of pTTS12 in conferring solvent tolerance. To this end, we cured the megaplasmid from P. putida S12 and conclusively confirmed that the SrpABC efflux pump is the major determinant of solvent tolerance on the megaplasmid pTTS12. In addition, we identified a novel toxin-antitoxin module (proposed gene names slvT and slvA, respectively) encoded on pTTS12 which contributes to the solvent tolerance phenotype and is important for conferring stability to the megaplasmid. Chromosomal introduction of the srp operon in combination with the slvAT gene pair created a solvent tolerance phenotype in non-solvent-tolerant strains, such as P. putida KT2440, Escherichia coli TG1, and E. coli BL21(DE3). IMPORTANCE Sustainable alternatives for high-value chemicals can be achieved by using renewable feedstocks in bacterial biocatalysis. However, during the bioproduction of such chemicals and biopolymers, aromatic compounds that function as products, substrates, or intermediates in the production process may exert toxicity to microbial host cells and limit the production yield. Therefore, solvent tolerance is a highly preferable trait for microbial hosts in the biobased production of aromatic chemicals and biopolymers. In this study, we revisit the essential role of megaplasmid pTTS12 from solvent-tolerant Pseudomonas putida S12 for molecular adaptation to an organic solvent. In addition to the solvent extrusion pump (SrpABC), we identified a novel toxin-antitoxin module (SlvAT) which contributes to short-term tolerance in moderate solvent concentrations, as well as to the stability of pTTS12. These two gene clusters were successfully expressed in non-solvent-tolerant strains of P. putida and Escherichia coli strains to confer and enhance solvent tolerance. American Society for Microbiology 2020-06-17 /pmc/articles/PMC7301842/ /pubmed/32358012 http://dx.doi.org/10.1128/AEM.00686-20 Text en Copyright © 2020 Kusumawardhani et al. https://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license (https://creativecommons.org/licenses/by/4.0/) .
spellingShingle Physiology
Kusumawardhani, Hadiastri
van Dijk, David
Hosseini, Rohola
de Winde, Johannes H.
Novel Toxin-Antitoxin Module SlvT-SlvA Regulates Megaplasmid Stability and Incites Solvent Tolerance in Pseudomonas putida S12
title Novel Toxin-Antitoxin Module SlvT-SlvA Regulates Megaplasmid Stability and Incites Solvent Tolerance in Pseudomonas putida S12
title_full Novel Toxin-Antitoxin Module SlvT-SlvA Regulates Megaplasmid Stability and Incites Solvent Tolerance in Pseudomonas putida S12
title_fullStr Novel Toxin-Antitoxin Module SlvT-SlvA Regulates Megaplasmid Stability and Incites Solvent Tolerance in Pseudomonas putida S12
title_full_unstemmed Novel Toxin-Antitoxin Module SlvT-SlvA Regulates Megaplasmid Stability and Incites Solvent Tolerance in Pseudomonas putida S12
title_short Novel Toxin-Antitoxin Module SlvT-SlvA Regulates Megaplasmid Stability and Incites Solvent Tolerance in Pseudomonas putida S12
title_sort novel toxin-antitoxin module slvt-slva regulates megaplasmid stability and incites solvent tolerance in pseudomonas putida s12
topic Physiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7301842/
https://www.ncbi.nlm.nih.gov/pubmed/32358012
http://dx.doi.org/10.1128/AEM.00686-20
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