L-天冬氨酸作为一种大宗氨基酸产品，在食品、医药和化工等方面有着广泛的用途。该研究在删除了富马酸水合酶基因(fumAC)的E. coli BL21 (DE3)中，构建了6种不同的马来酸顺反异构酶基因(maiA)与L-天冬氨酸裂解酶基因(aspA)双酶偶联的表达体系，通过比较其共表达效果和催化效率，获得最佳的偶联体系E. coli BL21 (DE3) ΔfumAC/pRSFDuet-1-maiA-aspA。该菌株能够在细胞浓度OD600值为8的催化体系下，在390 min内将3.2 mol/L的马来酸完全转化为L-天冬氨酸铵，其浓度达到3.14 mol/L，中间产物富马酸几乎没有积累，转化率达到98%以上。此外，为解决在高浓度的黏稠反应液中细胞回收利用困难的问题，对重组工程菌进行细胞固定化，固定化细胞回收利用8次之后，其相对酶活还剩下81%，显著提高了细胞的利用率，降低了L-天冬氨酸生产成本，为其工业化生产奠定了基础。

As a staple amino acid product, L-aspartic acid has been widely used in food, medicine and chemical industries. Six different maleate cis-trans isomerase and L-aspartic acid amino lyase co-expression systems have been built in E. coli BL21 (DE3) ΔfumAC in this study. By comparing their co-expression results and catalytic efficiency, the optimal co-expression system E. coli BL21 (DE3) ΔfumAC/pRSFDuet-1-maiA-aspA was obtained. 3.2 mol/L of maleic acid was completely transformed into L-aspartic acid ammonium in 390 minute with OD600 of 8. The concentration of L-aspartic acid ammonium reached 3.14 mol/L, conversion rate was above 98%, and the intermediate fumaric acid had almost no accumulation overall the reaction progress. Furthermore, to solve the problem that is difficult to recycle cells in high concentration of viscous reaction liquid, cell immobilization was studied. After 8 recycles of the immobilized cells, the relative enzyme activity was still 81%. The result suggested that the utilization rate of cells were greatly improved and the production cost reduced, which laid the foundation for the future industrial production of L-aspartic acid.