Engineering Escherichia coli for high-yielding 2,5-Dimethylpyrazine synthesis from L-Threonine by reconstructing metabolic pathways and enhancing cofactors regeneration

IF 6.1 1区工程技术 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Biotechnology for Biofuels Pub Date : 2024-03-18 DOI:10.1186/s13068-024-02487-4

Xin-Xin Liu, Yao Wang, Jian-Hui Zhang, Yun-Feng Lu, Zi-Xing Dong, Chao Yue, Xian-Qing Huang, Si-Pu Zhang, Dan-Dan Li, Lun-Guang Yao, Cun-Duo Tang

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Abstract

2,5-Dimethylpyrazine (2,5-DMP) is important pharmaceutical raw material and food flavoring agent. Recently, engineering microbes to produce 2,5-DMP has become an attractive alternative to chemical synthesis approach. In this study, metabolic engineering strategies were used to optimize the modified Escherichia coli BL21 (DE3) strain for efficient synthesis of 2,5-DMP using L-threonine dehydrogenase (EcTDH) from Escherichia coli BL21, NADH oxidase (EhNOX) from Enterococcus hirae, aminoacetone oxidase (ScAAO) from Streptococcus cristatus and L-threonine transporter protein (EcSstT) from Escherichia coli BL21, respectively. We further optimized the reaction conditions for synthesizing 2,5-DMP. In optimized conditions, the modified strain can convert L-threonine to obtain 2,5-DMP with a yield of 2897.30 mg/L. Therefore, the strategies used in this study contribute to the development of high-level cell factories for 2,5-DMP.

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通过重建新陈代谢途径和加强辅助因子再生，改造大肠杆菌，从 L-苏氨酸中高产合成 2,5-二甲基吡嗪。

2,5-二甲基吡嗪（2,5-DMP）是重要的医药原料和食品调味剂。最近，利用微生物工程技术生产 2,5-二甲基吡嗪已成为化学合成方法的一种有吸引力的替代方法。本研究采用代谢工程策略，分别利用大肠杆菌 BL21 的 L-苏氨酸脱氢酶（EcTDH）、平滑肠球菌的 NADH 氧化酶（EhNOX）、皱褶链球菌的氨基丙酮氧化酶（SCAAO）和大肠杆菌 BL21 的 L-苏氨酸转运蛋白（EcSstT），优化改造大肠杆菌 BL21（DE3）菌株，以高效合成 2,5-DMP。我们进一步优化了合成 2,5-DMP 的反应条件。在优化条件下，改良菌株可将 L-苏氨酸转化为 2,5-DMP，产率达 2897.30 mg/L。因此，本研究采用的策略有助于开发 2,5-二羟甲基丙烷的高级细胞工厂。

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来源期刊

Biotechnology for Biofuels 工程技术-生物工程与应用微生物

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审稿时长

2.7 months

期刊介绍： Biotechnology for Biofuels is an open access peer-reviewed journal featuring high-quality studies describing technological and operational advances in the production of biofuels, chemicals and other bioproducts. The journal emphasizes understanding and advancing the application of biotechnology and synergistic operations to improve plants and biological conversion systems for the biological production of these products from biomass, intermediates derived from biomass, or CO2, as well as upstream or downstream operations that are integral to biological conversion of biomass. Biotechnology for Biofuels focuses on the following areas: • Development of terrestrial plant feedstocks • Development of algal feedstocks • Biomass pretreatment, fractionation and extraction for biological conversion • Enzyme engineering, production and analysis • Bacterial genetics, physiology and metabolic engineering • Fungal/yeast genetics, physiology and metabolic engineering • Fermentation, biocatalytic conversion and reaction dynamics • Biological production of chemicals and bioproducts from biomass • Anaerobic digestion, biohydrogen and bioelectricity • Bioprocess integration, techno-economic analysis, modelling and policy • Life cycle assessment and environmental impact analysis