L-丝氨酸生产菌发酵及合成过程优化研究

Optimization of Fermention Conditions of A Strain for L-Serine Production and Its Synthesizing Process

【目的】研究并优化工程菌大肠杆菌(E.coli-SRZ018)的培养条件,确定四氢叶酸、磷酸吡哆醛和甲醛等对L-丝氨酸产量及转化率的影响。【方法】分别采用比浊法和液相-电喷雾质谱(HPLC-ESI/MS)测定菌体生物量、甘氨酸及L-丝氨酸含量。【结果】确定最优的培养基组成:磷酸氢二钾0.8 g/L、玉米浆35.0 g/L、甘油8.0 g/L、硫酸镁1.0 g/L、蛋白胨10.0 g/L,该条件下菌体生物量达2.88 g/L;进一步优化酶转化条件,即甘氨酸15.0 mmol/L、磷酸吡哆醛0.4 mmol/L、四氢叶酸7.0 mmol/L和甲醛10.0 mmol/L,维持反应体系p H 7.5,L-丝氨酸的转化率达95.6%。【结论】滴加甲醛能够显著提高酶法合成L-丝氨酸的底物转化率。

【Objective】Objective to optimize the medium components and culture condition of E.coliSRZ018,which is a kind of genetic engineering bacterium,observe the impact of tetrahydrofolate,pyridoxal phosphate,formaldehyde and other various factors on the conversion rates of substrate.【Method】Turbidimetric and HPLC-ESI/MS were used to determine biomass and amino acids respectively.【Result】The optimum composition of the medium was determined as follow:dipotassium phosphate 0.8 g/L,corn syrup 35 g/L,glycerol 8 g/L,magnesium sulfate 1 g/L,peptone 10 g/L.Consequently,the level of biomass reached 2.88 g/L.The influence of the critical parameters on substrate conversion rates was analyzed,and results showed the content of glycine,pyridoxal phosphate,tetrahydrofolate in enzyme-catalyzed solution were 15 mmol/L,0.4 mmol/L and 7 mmol/L respectively,and formaldehyde was added until its concentration reached 10 mmol/L,adjusting the pH of reaction system into 7.5,and conversion rate reached 95.6%.【Conclusion】Formol titration could improve the conversion rate of substrate significantly when enzymatic conversion technology for L-serine was applied.