Metabolic engineering of Bacillus amyloliquefaciens LL3 for enhanced poly‐γ‐glutamic acid synthesis
Summary Poly‐γ‐glutamic acid (γ‐PGA) is a biocompatible and biodegradable polypeptide with wide‐ranging applications in foods, cosmetics, medicine, agriculture and wastewater treatment. Bacillus amyloliquefaciens LL3 can produce γ‐PGA from sucrose that can be obtained easily from sugarcane and sugar...
| Main Authors: | , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Wiley
2019-09-01
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| Series: | Microbial Biotechnology |
| Online Access: | https://doi.org/10.1111/1751-7915.13446 |
| _version_ | 1818133639817330688 |
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| author | Weixia Gao Yulian He Fang Zhang Fengjie Zhao Chao Huang Yiting Zhang Qiang Zhao Shufang Wang Chao Yang |
| author_facet | Weixia Gao Yulian He Fang Zhang Fengjie Zhao Chao Huang Yiting Zhang Qiang Zhao Shufang Wang Chao Yang |
| author_sort | Weixia Gao |
| collection | DOAJ |
| description | Summary Poly‐γ‐glutamic acid (γ‐PGA) is a biocompatible and biodegradable polypeptide with wide‐ranging applications in foods, cosmetics, medicine, agriculture and wastewater treatment. Bacillus amyloliquefaciens LL3 can produce γ‐PGA from sucrose that can be obtained easily from sugarcane and sugar beet. In our previous work, it was found that low intracellular glutamate concentration was the limiting factor for γ‐PGA production by LL3. In this study, the γ‐PGA synthesis by strain LL3 was enhanced by chromosomally engineering its glutamate metabolism‐relevant networks. First, the downstream metabolic pathways were partly blocked by deleting fadR, lysC, aspB, pckA, proAB, rocG and gudB. The resulting strain NK‐A6 synthesized 4.84 g l−1 γ‐PGA, with a 31.5% increase compared with strain LL3. Second, a strong promoter PC2up was inserted into the upstream of icd gene, to generate strain NK‐A7, which further led to a 33.5% improvement in the γ‐PGA titre, achieving 6.46 g l−1. The NADPH level was improved by regulating the expression of pgi and gndA. Third, metabolic evolution was carried out to generate strain NK‐A9E, which showed a comparable γ‐PGA titre with strain NK‐A7. Finally, the srf and itu operons were deleted respectively, from the original strains NK‐A7 and NK‐A9E. The resulting strain NK‐A11 exhibited the highest γ‐PGA titre (7.53 g l−1), with a 2.05‐fold improvement compared with LL3. The results demonstrated that the approaches described here efficiently enhanced γ‐PGA production in B. amyloliquefaciens fermentation. |
| first_indexed | 2024-12-11T08:55:56Z |
| format | Article |
| id | doaj.art-4ab112cf016c453bb65f1b3f31082473 |
| institution | Directory Open Access Journal |
| issn | 1751-7915 |
| language | English |
| last_indexed | 2024-12-11T08:55:56Z |
| publishDate | 2019-09-01 |
| publisher | Wiley |
| record_format | Article |
| series | Microbial Biotechnology |
| spelling | doaj.art-4ab112cf016c453bb65f1b3f310824732022-12-22T01:13:53ZengWileyMicrobial Biotechnology1751-79152019-09-0112593294510.1111/1751-7915.13446Metabolic engineering of Bacillus amyloliquefaciens LL3 for enhanced poly‐γ‐glutamic acid synthesisWeixia Gao0Yulian He1Fang Zhang2Fengjie Zhao3Chao Huang4Yiting Zhang5Qiang Zhao6Shufang Wang7Chao Yang8Key Laboratory of Molecular Microbiology and Technology for Ministry of Education Nankai University Tianjin 300071 ChinaPrenatal Diagnosis and Genetic Diagnosis Center Tangshan Maternal and Child Health Care Hospital Tangshan 063000 ChinaKey Laboratory of Molecular Microbiology and Technology for Ministry of Education Nankai University Tianjin 300071 ChinaKey Laboratory of Molecular Microbiology and Technology for Ministry of Education Nankai University Tianjin 300071 ChinaKey Laboratory of Molecular Microbiology and Technology for Ministry of Education Nankai University Tianjin 300071 ChinaKey Laboratory of Molecular Microbiology and Technology for Ministry of Education Nankai University Tianjin 300071 ChinaState Key Laboratory of Medicinal Chemical Biology Nankai University Tianjin 300071 ChinaState Key Laboratory of Medicinal Chemical Biology Nankai University Tianjin 300071 ChinaKey Laboratory of Molecular Microbiology and Technology for Ministry of Education Nankai University Tianjin 300071 ChinaSummary Poly‐γ‐glutamic acid (γ‐PGA) is a biocompatible and biodegradable polypeptide with wide‐ranging applications in foods, cosmetics, medicine, agriculture and wastewater treatment. Bacillus amyloliquefaciens LL3 can produce γ‐PGA from sucrose that can be obtained easily from sugarcane and sugar beet. In our previous work, it was found that low intracellular glutamate concentration was the limiting factor for γ‐PGA production by LL3. In this study, the γ‐PGA synthesis by strain LL3 was enhanced by chromosomally engineering its glutamate metabolism‐relevant networks. First, the downstream metabolic pathways were partly blocked by deleting fadR, lysC, aspB, pckA, proAB, rocG and gudB. The resulting strain NK‐A6 synthesized 4.84 g l−1 γ‐PGA, with a 31.5% increase compared with strain LL3. Second, a strong promoter PC2up was inserted into the upstream of icd gene, to generate strain NK‐A7, which further led to a 33.5% improvement in the γ‐PGA titre, achieving 6.46 g l−1. The NADPH level was improved by regulating the expression of pgi and gndA. Third, metabolic evolution was carried out to generate strain NK‐A9E, which showed a comparable γ‐PGA titre with strain NK‐A7. Finally, the srf and itu operons were deleted respectively, from the original strains NK‐A7 and NK‐A9E. The resulting strain NK‐A11 exhibited the highest γ‐PGA titre (7.53 g l−1), with a 2.05‐fold improvement compared with LL3. The results demonstrated that the approaches described here efficiently enhanced γ‐PGA production in B. amyloliquefaciens fermentation.https://doi.org/10.1111/1751-7915.13446 |
| spellingShingle | Weixia Gao Yulian He Fang Zhang Fengjie Zhao Chao Huang Yiting Zhang Qiang Zhao Shufang Wang Chao Yang Metabolic engineering of Bacillus amyloliquefaciens LL3 for enhanced poly‐γ‐glutamic acid synthesis Microbial Biotechnology |
| title | Metabolic engineering of Bacillus amyloliquefaciens LL3 for enhanced poly‐γ‐glutamic acid synthesis |
| title_full | Metabolic engineering of Bacillus amyloliquefaciens LL3 for enhanced poly‐γ‐glutamic acid synthesis |
| title_fullStr | Metabolic engineering of Bacillus amyloliquefaciens LL3 for enhanced poly‐γ‐glutamic acid synthesis |
| title_full_unstemmed | Metabolic engineering of Bacillus amyloliquefaciens LL3 for enhanced poly‐γ‐glutamic acid synthesis |
| title_short | Metabolic engineering of Bacillus amyloliquefaciens LL3 for enhanced poly‐γ‐glutamic acid synthesis |
| title_sort | metabolic engineering of bacillus amyloliquefaciens ll3 for enhanced poly γ glutamic acid synthesis |
| url | https://doi.org/10.1111/1751-7915.13446 |
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