A temperature dependent pilin promoter for production of thermostable enzymes in Thermus thermophilus
Abstract Background Enzymes from thermophiles are of great interest for research and bioengineering due to their stability and efficiency. Thermophilic expression hosts such as Thermus thermophilus [T. thermophilus] can overcome specific challenges experienced with protein production in mesophilic e...
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| Format: | Article |
| Language: | English |
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BMC
2023-09-01
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| Series: | Microbial Cell Factories |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s12934-023-02192-1 |
| _version_ | 1797450989375062016 |
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| author | Lennart Kirchner Volker Müller Beate Averhoff |
| author_facet | Lennart Kirchner Volker Müller Beate Averhoff |
| author_sort | Lennart Kirchner |
| collection | DOAJ |
| description | Abstract Background Enzymes from thermophiles are of great interest for research and bioengineering due to their stability and efficiency. Thermophilic expression hosts such as Thermus thermophilus [T. thermophilus] can overcome specific challenges experienced with protein production in mesophilic expression hosts, such as leading to better folding, increased protein stability, solubility, and enzymatic activity. However, available inducible promoters for efficient protein production in T. thermophilus HB27 are limited. Results In this study, we characterized the pilA4 promoter region and evaluated its potential as a tool for production of thermostable enzymes in T. thermophilus HB27. Reporter gene analysis using a promoterless β-glucosidase gene revealed that the pilA4 promoter is highly active under optimal growth conditions at 68 °C and downregulated during growth at 80 °C. Furthermore, growth in minimal medium led to significantly increased promoter activity in comparison to growth in complex medium. Finally, we proved the suitability of the pilA4 promoter for heterologous production of thermostable enzymes in T. thermophilus by producing a fully active soluble mannitol-1-phosphate dehydrogenase from Thermoanaerobacter kivui [T. kivui], which is used in degradation of brown algae that are rich in mannitol. Conclusions Our results show that the pilA4 promoter is an efficient tool for gene expression in T. thermophilus with a high potential for use in biotechnology and synthetic biology applications. |
| first_indexed | 2024-03-09T14:48:28Z |
| format | Article |
| id | doaj.art-8ce8c9b3a36d456d8494fbc9bc45a1d7 |
| institution | Directory Open Access Journal |
| issn | 1475-2859 |
| language | English |
| last_indexed | 2024-03-09T14:48:28Z |
| publishDate | 2023-09-01 |
| publisher | BMC |
| record_format | Article |
| series | Microbial Cell Factories |
| spelling | doaj.art-8ce8c9b3a36d456d8494fbc9bc45a1d72023-11-26T14:37:49ZengBMCMicrobial Cell Factories1475-28592023-09-0122111010.1186/s12934-023-02192-1A temperature dependent pilin promoter for production of thermostable enzymes in Thermus thermophilusLennart Kirchner0Volker Müller1Beate Averhoff2Molecular Microbiology & Bioenergetics, Institute of Molecular Biosciences, Johann Wolfgang Goethe University Frankfurt/MainMolecular Microbiology & Bioenergetics, Institute of Molecular Biosciences, Johann Wolfgang Goethe University Frankfurt/MainMolecular Microbiology & Bioenergetics, Institute of Molecular Biosciences, Johann Wolfgang Goethe University Frankfurt/MainAbstract Background Enzymes from thermophiles are of great interest for research and bioengineering due to their stability and efficiency. Thermophilic expression hosts such as Thermus thermophilus [T. thermophilus] can overcome specific challenges experienced with protein production in mesophilic expression hosts, such as leading to better folding, increased protein stability, solubility, and enzymatic activity. However, available inducible promoters for efficient protein production in T. thermophilus HB27 are limited. Results In this study, we characterized the pilA4 promoter region and evaluated its potential as a tool for production of thermostable enzymes in T. thermophilus HB27. Reporter gene analysis using a promoterless β-glucosidase gene revealed that the pilA4 promoter is highly active under optimal growth conditions at 68 °C and downregulated during growth at 80 °C. Furthermore, growth in minimal medium led to significantly increased promoter activity in comparison to growth in complex medium. Finally, we proved the suitability of the pilA4 promoter for heterologous production of thermostable enzymes in T. thermophilus by producing a fully active soluble mannitol-1-phosphate dehydrogenase from Thermoanaerobacter kivui [T. kivui], which is used in degradation of brown algae that are rich in mannitol. Conclusions Our results show that the pilA4 promoter is an efficient tool for gene expression in T. thermophilus with a high potential for use in biotechnology and synthetic biology applications.https://doi.org/10.1186/s12934-023-02192-1Gene expressionThermophilesPromotersThermus thermophilus HB27Thermozymes |
| spellingShingle | Lennart Kirchner Volker Müller Beate Averhoff A temperature dependent pilin promoter for production of thermostable enzymes in Thermus thermophilus Microbial Cell Factories Gene expression Thermophiles Promoters Thermus thermophilus HB27 Thermozymes |
| title | A temperature dependent pilin promoter for production of thermostable enzymes in Thermus thermophilus |
| title_full | A temperature dependent pilin promoter for production of thermostable enzymes in Thermus thermophilus |
| title_fullStr | A temperature dependent pilin promoter for production of thermostable enzymes in Thermus thermophilus |
| title_full_unstemmed | A temperature dependent pilin promoter for production of thermostable enzymes in Thermus thermophilus |
| title_short | A temperature dependent pilin promoter for production of thermostable enzymes in Thermus thermophilus |
| title_sort | temperature dependent pilin promoter for production of thermostable enzymes in thermus thermophilus |
| topic | Gene expression Thermophiles Promoters Thermus thermophilus HB27 Thermozymes |
| url | https://doi.org/10.1186/s12934-023-02192-1 |
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