The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 1800# Lihu Road, Wuxi, 214122, China.
State Key Laboratory of Food Science and Technology, School of Biotechnology, Jiangnan University, 1800# Lihu Road, Wuxi, 214122, China.
Microb Cell Fact. 2020 Feb 18;19(1):39. doi: 10.1186/s12934-020-1294-7.
The efficiency of industrial fermentation process mainly depends on carbon yield, final titer and productivity. To improve the efficiency of L-lysine production from mixed sugar, we engineered carbohydrate metabolism systems to enhance the effective use of sugar in this study. A functional metabolic pathway of sucrose and fructose was engineered through introduction of fructokinase from Clostridium acetobutylicum. L-lysine production was further increased through replacement of phosphoenolpyruvate-dependent glucose and fructose uptake system (PTS and PTS) by inositol permeases (IolT1 and IolT2) and ATP-dependent glucokinase (ATP-GlK). However, the shortage of intracellular ATP has a significantly negative impact on sugar consumption rate, cell growth and L-lysine production. To overcome this defect, the recombinant strain was modified to co-express bifunctional ADP-dependent glucokinase (ADP-GlK/PFK) and NADH dehydrogenase (NDH-2) as well as to inactivate SigmaH factor (SigH), thus reducing the consumption of ATP and increasing ATP regeneration. Combination of these genetic modifications resulted in an engineered C. glutamicum strain K-8 capable of producing 221.3 ± 17.6 g/L L-lysine with productivity of 5.53 g/L/h and carbon yield of 0.71 g/g glucose in fed-batch fermentation. As far as we know, this is the best efficiency of L-lysine production from mixed sugar. This is also the first report for improving the efficiency of L-lysine production by systematic modification of carbohydrate metabolism systems.
工业发酵过程的效率主要取决于碳产率、最终浓度和生产力。为了提高混合糖生产 L-赖氨酸的效率，我们对碳水化合物代谢系统进行了工程改造，以增强糖的有效利用。通过引入丙酮丁醇梭菌的果糖激酶，构建了蔗糖和果糖的功能性代谢途径。通过肌醇通透酶（IolT1 和 IolT2）和 ATP 依赖性葡萄糖激酶（ATP-GlK）替代磷酸烯醇丙酮酸依赖性葡萄糖和果糖摄取系统（PTS 和 PTS），进一步提高了 L-赖氨酸的产量。然而，细胞内 ATP 的短缺对糖消耗速率、细胞生长和 L-赖氨酸的生产有显著的负面影响。为了克服这一缺陷，对重组菌株进行了修饰，共表达双功能 ADP 依赖性葡萄糖激酶（ADP-GlK/PFK）和 NADH 脱氢酶（NDH-2），并使 SigmaH 因子（SigH）失活，从而减少 ATP 的消耗并增加 ATP 的再生。这些遗传修饰的结合使工程化的 C. glutamicum 菌株 K-8 能够在分批补料发酵中生产 221.3 ± 17.6 g/L 的 L-赖氨酸，生产强度为 5.53 g/L/h，碳产率为 0.71 g/g 葡萄糖。据我们所知，这是混合糖生产 L-赖氨酸的最佳效率。这也是首次通过系统修饰碳水化合物代谢系统来提高 L-赖氨酸生产效率的报道。
