Engineering of l-threonine and l-proline biosensors by directed evolution of transcriptional regulator SerR and application for high-throughput screening
Abstract Amino acids are important bio-based products with a multi-billion-dollar market. The development of efficient high-throughput screening technologies utilizing biosensors is essential for the rapid identification of high-performance amino acid producers. However, there remains a pressing nee...
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| Format: | Article |
| Language: | English |
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SpringerOpen
2025-01-01
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| Series: | Bioresources and Bioprocessing |
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| Online Access: | https://doi.org/10.1186/s40643-024-00837-6 |
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| author | Wei Pu Jinhui Feng Jiuzhou Chen Jiao Liu Xuan Guo Lixian Wang Xiaojia Zhao Ningyun Cai Wenjuan Zhou Yu Wang Ping Zheng Jibin Sun |
| author_facet | Wei Pu Jinhui Feng Jiuzhou Chen Jiao Liu Xuan Guo Lixian Wang Xiaojia Zhao Ningyun Cai Wenjuan Zhou Yu Wang Ping Zheng Jibin Sun |
| author_sort | Wei Pu |
| collection | DOAJ |
| description | Abstract Amino acids are important bio-based products with a multi-billion-dollar market. The development of efficient high-throughput screening technologies utilizing biosensors is essential for the rapid identification of high-performance amino acid producers. However, there remains a pressing need for biosensors that specifically target certain critical amino acids, such as l-threonine and l-proline. In this study, a novel transcriptional regulator-based biosensor for l-threonine and l-proline was successfully developed, inspired by our new finding that SerE can export l-proline in addition to the previously known l-threonine and l-serine. Through directed evolution of SerR (the corresponding transcriptional regulator of SerE), the mutant SerRF104I which can recognize both l-threonine and l-proline as effectors and effectively distinguish strains with varying production levels was identified. Subsequently, the SerRF104I-based biosensor was employed for high-throughput screening of the superior enzyme mutants of l-homoserine dehydrogenase and γ-glutamyl kinase, which are critical enzymes in the biosynthesis of l-threonine and l-proline, respectively. A total of 25 and 13 novel mutants that increased the titers of l-threonine and l-proline by over 10% were successfully identified. Notably, six of the newly identified mutants exhibited similarities to the most effective mutants reported to date, indicating the promising application potential of the SerRF104I-based biosensor. This study illustrates an effective strategy for the development of transcriptional regulator-based biosensors for amino acids and other chemical compounds. |
| format | Article |
| id | doaj-art-2c88a483a927432586b66cf1179984a5 |
| institution | Kabale University |
| issn | 2197-4365 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | SpringerOpen |
| record_format | Article |
| series | Bioresources and Bioprocessing |
| spelling | doaj-art-2c88a483a927432586b66cf1179984a52025-01-19T12:07:53ZengSpringerOpenBioresources and Bioprocessing2197-43652025-01-0112111410.1186/s40643-024-00837-6Engineering of l-threonine and l-proline biosensors by directed evolution of transcriptional regulator SerR and application for high-throughput screeningWei Pu0Jinhui Feng1Jiuzhou Chen2Jiao Liu3Xuan Guo4Lixian Wang5Xiaojia Zhao6Ningyun Cai7Wenjuan Zhou8Yu Wang9Ping Zheng10Jibin Sun11Key Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesKey Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesKey Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesKey Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesKey Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesKey Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesKey Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesCollege of Biotechnology, Tianjin University of Science and TechnologyKey Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesKey Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesKey Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesKey Laboratory of Engineering Biology for Low-carbon Manufacturing, Tianjin Institute of Industrial Biotechnology, Chinese Academy of SciencesAbstract Amino acids are important bio-based products with a multi-billion-dollar market. The development of efficient high-throughput screening technologies utilizing biosensors is essential for the rapid identification of high-performance amino acid producers. However, there remains a pressing need for biosensors that specifically target certain critical amino acids, such as l-threonine and l-proline. In this study, a novel transcriptional regulator-based biosensor for l-threonine and l-proline was successfully developed, inspired by our new finding that SerE can export l-proline in addition to the previously known l-threonine and l-serine. Through directed evolution of SerR (the corresponding transcriptional regulator of SerE), the mutant SerRF104I which can recognize both l-threonine and l-proline as effectors and effectively distinguish strains with varying production levels was identified. Subsequently, the SerRF104I-based biosensor was employed for high-throughput screening of the superior enzyme mutants of l-homoserine dehydrogenase and γ-glutamyl kinase, which are critical enzymes in the biosynthesis of l-threonine and l-proline, respectively. A total of 25 and 13 novel mutants that increased the titers of l-threonine and l-proline by over 10% were successfully identified. Notably, six of the newly identified mutants exhibited similarities to the most effective mutants reported to date, indicating the promising application potential of the SerRF104I-based biosensor. This study illustrates an effective strategy for the development of transcriptional regulator-based biosensors for amino acids and other chemical compounds.https://doi.org/10.1186/s40643-024-00837-6l-Threonine and l-proline biosensorTranscriptional regulator SerRDirected evolutionHigh-throughput screeningl-Homoserine dehydrogenaseγ-Glutamyl kinase |
| spellingShingle | Wei Pu Jinhui Feng Jiuzhou Chen Jiao Liu Xuan Guo Lixian Wang Xiaojia Zhao Ningyun Cai Wenjuan Zhou Yu Wang Ping Zheng Jibin Sun Engineering of l-threonine and l-proline biosensors by directed evolution of transcriptional regulator SerR and application for high-throughput screening Bioresources and Bioprocessing l-Threonine and l-proline biosensor Transcriptional regulator SerR Directed evolution High-throughput screening l-Homoserine dehydrogenase γ-Glutamyl kinase |
| title | Engineering of l-threonine and l-proline biosensors by directed evolution of transcriptional regulator SerR and application for high-throughput screening |
| title_full | Engineering of l-threonine and l-proline biosensors by directed evolution of transcriptional regulator SerR and application for high-throughput screening |
| title_fullStr | Engineering of l-threonine and l-proline biosensors by directed evolution of transcriptional regulator SerR and application for high-throughput screening |
| title_full_unstemmed | Engineering of l-threonine and l-proline biosensors by directed evolution of transcriptional regulator SerR and application for high-throughput screening |
| title_short | Engineering of l-threonine and l-proline biosensors by directed evolution of transcriptional regulator SerR and application for high-throughput screening |
| title_sort | engineering of l threonine and l proline biosensors by directed evolution of transcriptional regulator serr and application for high throughput screening |
| topic | l-Threonine and l-proline biosensor Transcriptional regulator SerR Directed evolution High-throughput screening l-Homoserine dehydrogenase γ-Glutamyl kinase |
| url | https://doi.org/10.1186/s40643-024-00837-6 |
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