期刊
SCIENCE ADVANCES
卷 6, 期 21, 页码 -出版社
AMER ASSOC ADVANCEMENT SCIENCE
DOI: 10.1126/sciadv.aba2383
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资金
- National Key Research and Development Program of China [2018YFA0900300]
- National Natural Science Foundation of China [31870066]
- China Postdoctoral Science Foundation [2016 T90421]
- Fundamental Research Funds for the Central Universities [JUSRP51708A]
- Priority Academic Program Development of Jiangsu Higher Education Institutions
- 111 Project [111-2-06]
- National First-Class Discipline Program of Light Industry Technology and Engineering [LITE2018-06]
- Key Research and Development Program of Ningxia Hui Autonomous Region [2019BCH01002]
- Jiangsu province Collaborative Innovation Center for Modern Industrial Fermentation industry development program
- science and technology innovation team foundation of Ningxia Hui Autonomous Region [KJT2017001]
Trans-4-hydroxy-L-proline is produced by trans-proline-4-hydroxylase with L-proline through glucose fermentation. Here, we designed a thorough from A to Z strategy to significantly improve trans-4-hydroxy-L-proline production. Through rare codon selected evolution, Escherichia coli M1 produced 18.2 g L-1 L-proline. Metabolically engineered M6 with the deletion of putA, proP, putP, and aceA, and proB mutation focused carbon flux to L-proline and released its feedback inhibition. It produced 15.7 g L-1 trans-4-hydroxy-L-proline with 10 g L-1 L-proline retained. Furthermore, a tunable circuit based on quorum sensing attenuated L-proline hydroxylation flux, resulting in 43.2 g L-1 trans-4-hydroxy-L-proline with 4.3 g L-1 L-proline retained. Finally, rationally designed L-proline hydroxylase gave 54.8 g L-1 trans-4-hydroxy-L-proline in 60 hours almost without L-proline remaining-the highest production to date. The de novo engineering carbon flux through rare codon selected evolution, dynamic precursor modulation, and metabolic engineering provides a good technological platform for efficient hydroxyl amino acid synthesis.
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