Integration of Genetic and Process Engineering for Optimized Rhamnolipid Production Using Pseudomonas putida
Rhamnolipids are biosurfactants produced by microorganisms with the potential to replace synthetic compounds with petrochemical origin. To promote industrial use of rhamnolipids, recombinant rhamnolipid production from sugars needs to be intensified. Since this remains challenging, the aim of the pr...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2020-08-01
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| Series: | Frontiers in Bioengineering and Biotechnology |
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| Online Access: | https://www.frontiersin.org/article/10.3389/fbioe.2020.00976/full |
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| author | Till Tiso Till Tiso Nina Ihling Nina Ihling Sonja Kubicki Sonja Kubicki Andreas Biselli Andreas Biselli Andreas Schonhoff Andreas Schonhoff Isabel Bator Isabel Bator Stephan Thies Stephan Thies Tobias Karmainski Tobias Karmainski Sebastian Kruth Sebastian Kruth Anna-Lena Willenbrink Anna-Lena Willenbrink Anita Loeschcke Anita Loeschcke Petra Zapp Petra Zapp Andreas Jupke Andreas Jupke Karl-Erich Jaeger Karl-Erich Jaeger Karl-Erich Jaeger Jochen Büchs Jochen Büchs Lars M. Blank Lars M. Blank |
| author_facet | Till Tiso Till Tiso Nina Ihling Nina Ihling Sonja Kubicki Sonja Kubicki Andreas Biselli Andreas Biselli Andreas Schonhoff Andreas Schonhoff Isabel Bator Isabel Bator Stephan Thies Stephan Thies Tobias Karmainski Tobias Karmainski Sebastian Kruth Sebastian Kruth Anna-Lena Willenbrink Anna-Lena Willenbrink Anita Loeschcke Anita Loeschcke Petra Zapp Petra Zapp Andreas Jupke Andreas Jupke Karl-Erich Jaeger Karl-Erich Jaeger Karl-Erich Jaeger Jochen Büchs Jochen Büchs Lars M. Blank Lars M. Blank |
| author_sort | Till Tiso |
| collection | DOAJ |
| description | Rhamnolipids are biosurfactants produced by microorganisms with the potential to replace synthetic compounds with petrochemical origin. To promote industrial use of rhamnolipids, recombinant rhamnolipid production from sugars needs to be intensified. Since this remains challenging, the aim of the presented research is to utilize a multidisciplinary approach to take a step toward developing a sustainable rhamnolipid production process. Here, we developed expression cassettes for stable integration of the rhamnolipid biosynthesis genes into the genome outperformed plasmid-based expression systems. Furthermore, the genetic stability of the production strain was improved by using an inducible promoter. To enhance rhamnolipid synthesis, energy- and/or carbon-consuming traits were removed: mutants negative for the synthesis of the flagellar machinery or the storage polymer PHA showed increased production by 50%. Variation of time of induction resulted in an 18% increase in titers. A scale-up from shake flasks was carried out using a 1-L bioreactor. By recycling of the foam, biomass loss could be minimized and a rhamnolipid titer of up to 1.5 g/L was achieved without using mechanical foam destroyers or antifoaming agents. Subsequent liquid–liquid extraction was optimized by using a suitable minimal medium during fermentation to reduce undesired interphase formation. A technical-scale production process was designed and evaluated by a life-cycle assessment (LCA). Different process chains and their specific environmental impact were examined. It was found that next to biomass supply, the fermentation had the biggest environmental impact. The present work underlines the need for multidisciplinary approaches to address the challenges associated with achieving sustainable production of microbial secondary metabolites. The results are discussed in the context of the challenges of microbial biosurfactant production using hydrophilic substrates on an industrial scale. |
| first_indexed | 2024-12-19T00:21:37Z |
| format | Article |
| id | doaj.art-4fecae5c422c41a3a7664b1484539c53 |
| institution | Directory Open Access Journal |
| issn | 2296-4185 |
| language | English |
| last_indexed | 2024-12-19T00:21:37Z |
| publishDate | 2020-08-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Bioengineering and Biotechnology |
| spelling | doaj.art-4fecae5c422c41a3a7664b1484539c532022-12-21T20:45:31ZengFrontiers Media S.A.Frontiers in Bioengineering and Biotechnology2296-41852020-08-01810.3389/fbioe.2020.00976565499Integration of Genetic and Process Engineering for Optimized Rhamnolipid Production Using Pseudomonas putidaTill Tiso0Till Tiso1Nina Ihling2Nina Ihling3Sonja Kubicki4Sonja Kubicki5Andreas Biselli6Andreas Biselli7Andreas Schonhoff8Andreas Schonhoff9Isabel Bator10Isabel Bator11Stephan Thies12Stephan Thies13Tobias Karmainski14Tobias Karmainski15Sebastian Kruth16Sebastian Kruth17Anna-Lena Willenbrink18Anna-Lena Willenbrink19Anita Loeschcke20Anita Loeschcke21Petra Zapp22Petra Zapp23Andreas Jupke24Andreas Jupke25Karl-Erich Jaeger26Karl-Erich Jaeger27Karl-Erich Jaeger28Jochen Büchs29Jochen Büchs30Lars M. Blank31Lars M. Blank32iAMB – Institute of Applied Microbiology, ABBt – Aachen Biology and Biotechnology, RWTH Aachen University, Aachen, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyChair of Biochemical Engineering (AVT.BioVT), RWTH Aachen University, Aachen, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyInstitute of Molecular Enzyme Technology, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyFluid Process Engineering (AVT.FVT), RWTH Aachen University, Aachen, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyInstitute of Energy and Climate Research – Systems Analysis and Technology Evaluation (IEK-STE), Forschungszentrum Jülich GmbH, Jülich, GermanyiAMB – Institute of Applied Microbiology, ABBt – Aachen Biology and Biotechnology, RWTH Aachen University, Aachen, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyInstitute of Molecular Enzyme Technology, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, GermanyiAMB – Institute of Applied Microbiology, ABBt – Aachen Biology and Biotechnology, RWTH Aachen University, Aachen, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyiAMB – Institute of Applied Microbiology, ABBt – Aachen Biology and Biotechnology, RWTH Aachen University, Aachen, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyFluid Process Engineering (AVT.FVT), RWTH Aachen University, Aachen, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyInstitute of Molecular Enzyme Technology, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyInstitute of Energy and Climate Research – Systems Analysis and Technology Evaluation (IEK-STE), Forschungszentrum Jülich GmbH, Jülich, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyFluid Process Engineering (AVT.FVT), RWTH Aachen University, Aachen, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyInstitute of Molecular Enzyme Technology, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, GermanyInstitute of Bio- and Geosciences IBG 1: Biotechnology, Forschungszentrum Jülich GmbH, Jülich, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyChair of Biochemical Engineering (AVT.BioVT), RWTH Aachen University, Aachen, GermanyiAMB – Institute of Applied Microbiology, ABBt – Aachen Biology and Biotechnology, RWTH Aachen University, Aachen, GermanyBioeconomy Science Center (BioSC), Forschungszentrum Jülich GmbH, Jülich, GermanyRhamnolipids are biosurfactants produced by microorganisms with the potential to replace synthetic compounds with petrochemical origin. To promote industrial use of rhamnolipids, recombinant rhamnolipid production from sugars needs to be intensified. Since this remains challenging, the aim of the presented research is to utilize a multidisciplinary approach to take a step toward developing a sustainable rhamnolipid production process. Here, we developed expression cassettes for stable integration of the rhamnolipid biosynthesis genes into the genome outperformed plasmid-based expression systems. Furthermore, the genetic stability of the production strain was improved by using an inducible promoter. To enhance rhamnolipid synthesis, energy- and/or carbon-consuming traits were removed: mutants negative for the synthesis of the flagellar machinery or the storage polymer PHA showed increased production by 50%. Variation of time of induction resulted in an 18% increase in titers. A scale-up from shake flasks was carried out using a 1-L bioreactor. By recycling of the foam, biomass loss could be minimized and a rhamnolipid titer of up to 1.5 g/L was achieved without using mechanical foam destroyers or antifoaming agents. Subsequent liquid–liquid extraction was optimized by using a suitable minimal medium during fermentation to reduce undesired interphase formation. A technical-scale production process was designed and evaluated by a life-cycle assessment (LCA). Different process chains and their specific environmental impact were examined. It was found that next to biomass supply, the fermentation had the biggest environmental impact. The present work underlines the need for multidisciplinary approaches to address the challenges associated with achieving sustainable production of microbial secondary metabolites. The results are discussed in the context of the challenges of microbial biosurfactant production using hydrophilic substrates on an industrial scale.https://www.frontiersin.org/article/10.3389/fbioe.2020.00976/fullrhamnolipidsPseudomonas putida KT2440synthetic biologymetabolic engineeringoxygen transfer rateliquid–liquid extraction |
| spellingShingle | Till Tiso Till Tiso Nina Ihling Nina Ihling Sonja Kubicki Sonja Kubicki Andreas Biselli Andreas Biselli Andreas Schonhoff Andreas Schonhoff Isabel Bator Isabel Bator Stephan Thies Stephan Thies Tobias Karmainski Tobias Karmainski Sebastian Kruth Sebastian Kruth Anna-Lena Willenbrink Anna-Lena Willenbrink Anita Loeschcke Anita Loeschcke Petra Zapp Petra Zapp Andreas Jupke Andreas Jupke Karl-Erich Jaeger Karl-Erich Jaeger Karl-Erich Jaeger Jochen Büchs Jochen Büchs Lars M. Blank Lars M. Blank Integration of Genetic and Process Engineering for Optimized Rhamnolipid Production Using Pseudomonas putida Frontiers in Bioengineering and Biotechnology rhamnolipids Pseudomonas putida KT2440 synthetic biology metabolic engineering oxygen transfer rate liquid–liquid extraction |
| title | Integration of Genetic and Process Engineering for Optimized Rhamnolipid Production Using Pseudomonas putida |
| title_full | Integration of Genetic and Process Engineering for Optimized Rhamnolipid Production Using Pseudomonas putida |
| title_fullStr | Integration of Genetic and Process Engineering for Optimized Rhamnolipid Production Using Pseudomonas putida |
| title_full_unstemmed | Integration of Genetic and Process Engineering for Optimized Rhamnolipid Production Using Pseudomonas putida |
| title_short | Integration of Genetic and Process Engineering for Optimized Rhamnolipid Production Using Pseudomonas putida |
| title_sort | integration of genetic and process engineering for optimized rhamnolipid production using pseudomonas putida |
| topic | rhamnolipids Pseudomonas putida KT2440 synthetic biology metabolic engineering oxygen transfer rate liquid–liquid extraction |
| url | https://www.frontiersin.org/article/10.3389/fbioe.2020.00976/full |
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