A Plasmid-Based Fluorescence Reporter System for Monitoring Oxidative Damage in <i>E. coli</i>
Quantitating intracellular oxidative damage caused by reactive oxygen species (ROS) is of interest in many fields of biological research. The current systems primarily rely on supplemented oxygen-sensitive substrates that penetrate the target cells, and react with ROS to produce signals that can be...
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| Format: | Article |
| Language: | English |
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MDPI AG
2022-08-01
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| Series: | Sensors |
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| Online Access: | https://www.mdpi.com/1424-8220/22/17/6334 |
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| author | Hariharan Dandapani Pasi Kankaanpää Patrik R. Jones Pauli Kallio |
| author_facet | Hariharan Dandapani Pasi Kankaanpää Patrik R. Jones Pauli Kallio |
| author_sort | Hariharan Dandapani |
| collection | DOAJ |
| description | Quantitating intracellular oxidative damage caused by reactive oxygen species (ROS) is of interest in many fields of biological research. The current systems primarily rely on supplemented oxygen-sensitive substrates that penetrate the target cells, and react with ROS to produce signals that can be monitored with spectroscopic or imaging techniques. The objective here was to design a new non-invasive analytical strategy for measuring ROS-induced damage inside living cells by taking advantage of the native redox sensor system of <i>E. coli</i>. The developed plasmid-based sensor relies on an oxygen-sensitive transcriptional repressor IscR that controls the expression of a fluorescent marker in vivo. The system was shown to quantitatively respond to oxidative stress induced by supplemented H<sub>2</sub>O<sub>2</sub> and lowered cultivation temperatures. Comparative analysis with fluorescence microscopy further demonstrated that the specificity of the reporter system was equivalent to the commercial chemical probe (CellROX). The strategy introduced here is not dependent on chemical probes, but instead uses a fluorescent expression system to detect enzyme-level oxidative damage in microbial cells. This provides a cheap and simple means for analysing enzyme-level oxidative damage in a biological context in <i>E. coli</i>. |
| first_indexed | 2024-03-10T01:17:13Z |
| format | Article |
| id | doaj.art-84b61ba1c989449e8828cfb8c5c0fab1 |
| institution | Directory Open Access Journal |
| issn | 1424-8220 |
| language | English |
| last_indexed | 2024-03-10T01:17:13Z |
| publishDate | 2022-08-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Sensors |
| spelling | doaj.art-84b61ba1c989449e8828cfb8c5c0fab12023-11-23T14:06:43ZengMDPI AGSensors1424-82202022-08-012217633410.3390/s22176334A Plasmid-Based Fluorescence Reporter System for Monitoring Oxidative Damage in <i>E. coli</i>Hariharan Dandapani0Pasi Kankaanpää1Patrik R. Jones2Pauli Kallio3Molecular Plant Biology, Department of Life Technologies, University of Turku, FI-20014 Turku, FinlandTurku BioImaging and Turku Bioscience Centre, University of Turku, FI-20014 Turku, FinlandMolecular Plant Biology, Department of Life Technologies, University of Turku, FI-20014 Turku, FinlandMolecular Plant Biology, Department of Life Technologies, University of Turku, FI-20014 Turku, FinlandQuantitating intracellular oxidative damage caused by reactive oxygen species (ROS) is of interest in many fields of biological research. The current systems primarily rely on supplemented oxygen-sensitive substrates that penetrate the target cells, and react with ROS to produce signals that can be monitored with spectroscopic or imaging techniques. The objective here was to design a new non-invasive analytical strategy for measuring ROS-induced damage inside living cells by taking advantage of the native redox sensor system of <i>E. coli</i>. The developed plasmid-based sensor relies on an oxygen-sensitive transcriptional repressor IscR that controls the expression of a fluorescent marker in vivo. The system was shown to quantitatively respond to oxidative stress induced by supplemented H<sub>2</sub>O<sub>2</sub> and lowered cultivation temperatures. Comparative analysis with fluorescence microscopy further demonstrated that the specificity of the reporter system was equivalent to the commercial chemical probe (CellROX). The strategy introduced here is not dependent on chemical probes, but instead uses a fluorescent expression system to detect enzyme-level oxidative damage in microbial cells. This provides a cheap and simple means for analysing enzyme-level oxidative damage in a biological context in <i>E. coli</i>.https://www.mdpi.com/1424-8220/22/17/6334stress responseoxidative stress damage sensoriron–sulphur clusterreactive oxygen specieslowered growth temperatureplasmid-based fluorescent reporter |
| spellingShingle | Hariharan Dandapani Pasi Kankaanpää Patrik R. Jones Pauli Kallio A Plasmid-Based Fluorescence Reporter System for Monitoring Oxidative Damage in <i>E. coli</i> Sensors stress response oxidative stress damage sensor iron–sulphur cluster reactive oxygen species lowered growth temperature plasmid-based fluorescent reporter |
| title | A Plasmid-Based Fluorescence Reporter System for Monitoring Oxidative Damage in <i>E. coli</i> |
| title_full | A Plasmid-Based Fluorescence Reporter System for Monitoring Oxidative Damage in <i>E. coli</i> |
| title_fullStr | A Plasmid-Based Fluorescence Reporter System for Monitoring Oxidative Damage in <i>E. coli</i> |
| title_full_unstemmed | A Plasmid-Based Fluorescence Reporter System for Monitoring Oxidative Damage in <i>E. coli</i> |
| title_short | A Plasmid-Based Fluorescence Reporter System for Monitoring Oxidative Damage in <i>E. coli</i> |
| title_sort | plasmid based fluorescence reporter system for monitoring oxidative damage in i e coli i |
| topic | stress response oxidative stress damage sensor iron–sulphur cluster reactive oxygen species lowered growth temperature plasmid-based fluorescent reporter |
| url | https://www.mdpi.com/1424-8220/22/17/6334 |
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