Liu Rongming, Ma Jiangfeng, Liang Liya, Xu Bing, Wang Guangming, Zhang Min, Jiang Min
State Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing University of Technology, Nanjing 210009, China.
Sheng Wu Gong Cheng Xue Bao. 2011 Oct;27(10):1438-47.
Escherichia coli strain NZN111 is a promising candidate for the fermentative production of succinate. However, because lactate dehydrogenase and pyruvate formate lyase were inactivated in NZN111, this strain had an unbalanced NADH/NAD+ ratio and could not use glucose under anaerobic conditions. In this study, a recombinant strain E. coli NZN111/pTrc99a-pncB was constructed to overexpress the nicotinic acid phosphoribosyl transferase gene (pncB). Under anaerobic conditions with the addition of 0.5 mmol/L nicotinic acid and 0.3 mmol/L isopropyl beta-D-thiogalactopyranoside (IPTG), the specific nicotinic acid phosphoribosyl transferase (NAPRTase, EC 2.4.2.11) activity in the recombinant strain was 11-fold higher than that in E. coli NZN111, the concentration of NAD(H) was increased by 3.85-fold, especially the concentration of NAD+ was increased by 5.17-fold and NADH/NAD+ was decreased from 0.640 to 0.125. The recombinant strain regained the capability of growth and glucose utilization under anaerobic conditions.
大肠杆菌菌株NZN111是发酵生产琥珀酸的一个有潜力的候选菌株。然而，由于NZN111中的乳酸脱氢酶和丙酮酸甲酸裂解酶被灭活，该菌株的NADH/NAD⁺比例失衡，在厌氧条件下无法利用葡萄糖。在本研究中，构建了重组菌株大肠杆菌NZN111/pTrc99a-pncB以过量表达烟酸磷酸核糖基转移酶基因（pncB）。在添加0.5 mmol/L烟酸和0.3 mmol/L异丙基-β-D-硫代半乳糖苷（IPTG）的厌氧条件下，重组菌株中特异性烟酸磷酸核糖基转移酶（NAPRTase，EC 2.4.2.11）的活性比大肠杆菌NZN111高11倍，NAD(H)的浓度增加了3.85倍，尤其是NAD⁺的浓度增加了5.17倍，NADH/NAD⁺从0.640降至0.125。该重组菌株在厌氧条件下恢复了生长和利用葡萄糖的能力。
Zotero 插件