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Real-Time Monitoring of the Yeast Intracellular State During Bioprocesses With a Toolbox of Biosensors

Industrial fermentation processes strive for high robustness to ensure optimal and consistent performance. Medium components, fermentation products, and physical perturbations may cause stress and lower performance. Cellular stress elicits a range of responses, whose extracellular manifestations hav...

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Autores principales: Torello Pianale, Luca, Rugbjerg, Peter, Olsson, Lisbeth
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8776715/
https://www.ncbi.nlm.nih.gov/pubmed/35069506
http://dx.doi.org/10.3389/fmicb.2021.802169
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author Torello Pianale, Luca
Rugbjerg, Peter
Olsson, Lisbeth
author_facet Torello Pianale, Luca
Rugbjerg, Peter
Olsson, Lisbeth
author_sort Torello Pianale, Luca
collection PubMed
description Industrial fermentation processes strive for high robustness to ensure optimal and consistent performance. Medium components, fermentation products, and physical perturbations may cause stress and lower performance. Cellular stress elicits a range of responses, whose extracellular manifestations have been extensively studied; whereas intracellular aspects remain poorly known due to lack of tools for real-time monitoring. Genetically encoded biosensors have emerged as promising tools and have been used to improve microbial productivity and tolerance toward industrially relevant stresses. Here, fluorescent biosensors able to sense the yeast intracellular environment (pH, ATP levels, oxidative stress, glycolytic flux, and ribosome production) were implemented into a versatile and easy-to-use toolbox. Marker-free and efficient genome integration at a conserved site on chromosome X of Saccharomyces cerevisiae strains and a commercial Saccharomyces boulardii strain was developed. Moreover, multiple biosensors were used to simultaneously monitor different intracellular parameters in a single cell. Even when combined together, the biosensors did not significantly affect key physiological parameters, such as specific growth rate and product yields. Activation and response of each biosensor and their interconnection were assessed using an advanced micro-cultivation system. Finally, the toolbox was used to screen cell behavior in a synthetic lignocellulosic hydrolysate that mimicked harsh industrial substrates, revealing differences in the oxidative stress response between laboratory (CEN.PK113-7D) and industrial (Ethanol Red) S. cerevisiae strains. In summary, the toolbox will allow both the exploration of yeast diversity and physiological responses in natural and complex industrial conditions, as well as the possibility to monitor production processes.
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spelling pubmed-87767152022-01-22 Real-Time Monitoring of the Yeast Intracellular State During Bioprocesses With a Toolbox of Biosensors Torello Pianale, Luca Rugbjerg, Peter Olsson, Lisbeth Front Microbiol Microbiology Industrial fermentation processes strive for high robustness to ensure optimal and consistent performance. Medium components, fermentation products, and physical perturbations may cause stress and lower performance. Cellular stress elicits a range of responses, whose extracellular manifestations have been extensively studied; whereas intracellular aspects remain poorly known due to lack of tools for real-time monitoring. Genetically encoded biosensors have emerged as promising tools and have been used to improve microbial productivity and tolerance toward industrially relevant stresses. Here, fluorescent biosensors able to sense the yeast intracellular environment (pH, ATP levels, oxidative stress, glycolytic flux, and ribosome production) were implemented into a versatile and easy-to-use toolbox. Marker-free and efficient genome integration at a conserved site on chromosome X of Saccharomyces cerevisiae strains and a commercial Saccharomyces boulardii strain was developed. Moreover, multiple biosensors were used to simultaneously monitor different intracellular parameters in a single cell. Even when combined together, the biosensors did not significantly affect key physiological parameters, such as specific growth rate and product yields. Activation and response of each biosensor and their interconnection were assessed using an advanced micro-cultivation system. Finally, the toolbox was used to screen cell behavior in a synthetic lignocellulosic hydrolysate that mimicked harsh industrial substrates, revealing differences in the oxidative stress response between laboratory (CEN.PK113-7D) and industrial (Ethanol Red) S. cerevisiae strains. In summary, the toolbox will allow both the exploration of yeast diversity and physiological responses in natural and complex industrial conditions, as well as the possibility to monitor production processes. Frontiers Media S.A. 2022-01-07 /pmc/articles/PMC8776715/ /pubmed/35069506 http://dx.doi.org/10.3389/fmicb.2021.802169 Text en Copyright © 2022 Torello Pianale, Rugbjerg and Olsson. 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 Microbiology
Torello Pianale, Luca
Rugbjerg, Peter
Olsson, Lisbeth
Real-Time Monitoring of the Yeast Intracellular State During Bioprocesses With a Toolbox of Biosensors
title Real-Time Monitoring of the Yeast Intracellular State During Bioprocesses With a Toolbox of Biosensors
title_full Real-Time Monitoring of the Yeast Intracellular State During Bioprocesses With a Toolbox of Biosensors
title_fullStr Real-Time Monitoring of the Yeast Intracellular State During Bioprocesses With a Toolbox of Biosensors
title_full_unstemmed Real-Time Monitoring of the Yeast Intracellular State During Bioprocesses With a Toolbox of Biosensors
title_short Real-Time Monitoring of the Yeast Intracellular State During Bioprocesses With a Toolbox of Biosensors
title_sort real-time monitoring of the yeast intracellular state during bioprocesses with a toolbox of biosensors
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8776715/
https://www.ncbi.nlm.nih.gov/pubmed/35069506
http://dx.doi.org/10.3389/fmicb.2021.802169
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