Zeng Weizhu, Cai Wen, Liu Li, Du Guocheng, Chen Jian, Zhou Jingwen
Key Laboratory of Industrial Biotechnology, Ministry of Education and School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu, 214122, China.
National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu, 214122, China.
Synth Syst Biotechnol. 2019 Jul 27;4(3):134-141. doi: 10.1016/j.synbio.2019.07.001. eCollection 2019 Sep.
2-keto-d-gluconic acid (2-KGA) is a key precursor for synthesising vitamin C and isovitamin C. However, phage contamination is as constant problem in industrial production of 2-KGA using . holds promise for producing 2-KGA due to impressive resistance to hypertonicity and acids, and high utilisation of glucose. In this study, the 2-KGA synthesis pathway was regulated to enhance production of 2-KGA and reduce accumulation of the by-products 5-keto-d-gluconic acid (5-KGA) and d-gluconic acid (D-GA) in the 2-KGA producer CGMCC 1.49. Knocking out the gene from a competitive pathway and overexpressing the gene from the 2-KGA synthesis pathway via homologous recombination increased the titre of 2-KGA by 63.81% in shake flasks. Additionally, accumulation of 5-KGA was decreased by 63.52% with the resulting -Δ- strain. Using an intermittent fed-batch mode in a 3 L fermenter, 2-KGA reached 235.3 g L with a 91.1% glucose conversion rate. Scaling up in a 15 L fermenter led to stable 2-KGA titre with productivity of 2.99 g L h, 11.99% higher than in the 3 L fermenter, and D-GA and 5-KGA by-products were completely converted to 2-KGA.
2-酮基-D-葡萄糖酸（2-KGA）是合成维生素C和异维生素C的关键前体。然而，在利用[具体方法]进行2-KGA的工业生产中，噬菌体污染一直是个问题。[某种微生物]由于对高渗和酸具有令人印象深刻的抗性以及对葡萄糖的高利用率，有望用于生产2-KGA。在本研究中，对2-KGA合成途径进行调控，以提高2-KGA生产者CGMCC 1.49中2-KGA的产量，并减少副产物5-酮基-D-葡萄糖酸（5-KGA）和D-葡萄糖酸（D-GA）的积累。通过同源重组从竞争途径中敲除[某基因]并过表达2-KGA合成途径中的[某基因]，在摇瓶中使2-KGA的产量提高了63.81%。此外，所得的[缺失某基因的菌株名称]-Δ-[缺失某基因名称]菌株使5-KGA的积累减少了63.52%。在3 L发酵罐中采用间歇补料分批模式，2-KGA达到235.3 g/L，葡萄糖转化率为91.1%。在15 L发酵罐中放大培养导致2-KGA产量稳定，生产率为2.99 g/L·h，比3 L发酵罐中的生产率高11.99%，并且副产物D-GA和5-KGA完全转化为2-KGA。
