Efficient production of 2-keto-L-gulonic acid from D-glucose in Gluconobacter oxydans ATCC9937 by mining key enzyme and transporter.
Efficient production of 2-keto-L-gulonic acid from D-glucose in Gluconobacter oxydans ATCC9937 by mining key enzyme and transporter.
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DOI：
10.1016/j.biortech.2023.129316
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发表时间：
2023-06
影响因子：
11.4
通讯作者：
Guang-wei Li;Dong Li;Weizhu Zeng;Zhi-jie Qin;Jian Chen;Jingwen Zhou
中科院分区：
文献类型：
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作者：
Guang-wei Li;Dong Li;Weizhu Zeng;Zhi-jie Qin;Jian Chen;Jingwen Zhou
由D-葡萄糖经2，5-二酮基-D-葡萄糖酸（2，5-DKG）直接生产2-酮基-L-古洛糖酸（2-KLG）是一条很有前途的替代路线。为探索D-葡萄糖转化为2-KLG的途径，选择葡萄糖氧化酶ATCC 9937为基础菌株。发现该菌株具有天然的由D-葡萄糖合成2-KLG的能力，并在其基因组上发现了一个新的2，5-DKG还原酶（DKGR）。确定了限制生产的几个主要问题，包括DKGR的催化能力不足，2，5-DKG的跨膜运动差以及宿主菌株细胞内外D-葡萄糖消耗通量不平衡。通过鉴定新的DKGR和2，5-DKG转运蛋白，通过平衡细胞内和细胞外D-葡萄糖代谢通量，系统地增强了整个2-KLG生物合成途径。工程菌产2-KLG 30.5g/L，转化率为39.0%。该结果为更经济的维生素C大规模发酵工艺铺平了道路。
Direct production of 2-keto-L-gulonic acid (2-KLG, the precursor of vitamin C) from D-glucose through 2,5-diketo-D-gluconic acid (2,5-DKG) is a promising alternative route. To explore the pathway of producing 2-KLG from D-glucose,Gluconobacter oxydansATCC9937 was selected as a chassis strain. It was found that the chassis strain naturally has the ability to synthesize 2-KLG from D-glucose, and a new 2,5-DKG reductase (DKGR) was found on its genome. Several major issues limiting production were identified, including the insufficient catalytic capacity of DKGR, poor transmembrane movement of 2,5-DKG and imbalanced D-glucose consumption flux inside and outside of the host strain cells. By identifying novel DKGR and 2,5-DKG transporter, the whole 2-KLG biosynthesis pathway was systematically enhanced by balancing intracellular and extracellular D-glucose metabolic flux. The engineered strain produced 30.5 g/L 2-KLG with a conversion ratio of 39.0%. The results pave the way for a more economical large-scale fermentation process for vitamin C.