重组大肠杆菌利用蔗糖及糖蜜发酵生产丁二酸

Succinic acid production from sucrose and sugarcane molasses by metabolically engineered Escherichia coli

摘要:

富含蔗糖的甘蔗糖蜜可作为制备丁二酸的廉价原料。然而生产丁二酸的潜力菌株大肠杆菌Escherichia coli AFP111不能代谢蔗糖。为了使其具有蔗糖代谢能力，将E. coli W中非PTS蔗糖利用系统蔗糖通透酶的编码基因cscB，果糖激酶的编码基因cscK和蔗糖水解酶的编码基因cscA克隆并表达到AFP111中，获得重组菌株AFP111/pMD19T-cscBKA。经厌氧发酵验证，重组菌株72 h消耗20 g/L蔗糖，丁二酸产量达到12 g/L。在3 L发酵罐中采用有氧阶段培养菌体、厌氧阶段发酵的两阶段发酵方式，厌氧发酵30 h，重组菌株以蔗糖和糖蜜为碳源丁二酸产量分别为34 g/L和30 g/L。结果表明，通过外源引入非PTS蔗糖利用系统，重组菌株具有较强的代谢蔗糖生长及合成丁二酸的能力，并且能够利用廉价糖蜜发酵制备丁二酸。

Abstract:

Sugarcane molasses containing large amounts of sucrose is an economical substrate for succinic acid production. However, Escherichia coli AFP111 cannot metabolize sucrose although it is a promising candidate for succinic acid production. To achieve sucrose utilizing ability, we cloned and expressed cscBKA genes encoding sucrose permease, fructokinase and invertase of non-PTS sucrose-utilization system from E. coli W in E. coli AFP111 to generate a recombinant strain AFP111/pMD19T-cscBKA. After 72 h of anaerobic fermentation of the recombinant in serum bottles, 20 g/L sucrose was consumed and 12 g/L succinic acid was produced. During dual-phase fermentation comprised of initial aerobic growth phase followed by anaerobic fermentation phase, the concentration of succinic acid from sucrose and sugarcane molasses was 34 g/L and 30 g/L, respectively, at 30 h of anaerobic phase in a 3 L fermentor. The results show that the introduction of non-PTS sucrose-utilization system has sucrose-metabolizing capability for cell growth and succinic acid production, and can use cheap sugarcane molasses to produce succinic acid.