The functional rare sugar D-allulose is mainly produced from fructose by the catalyzation of D-allulose 3-epimerase (DAE) from genetically engineered microorganisms. In this study, a DAE expression recombinant Escherichia coli was chosen as the object, and single factor experiments were carried out to study the influence of carbon source, nitrogen source, and metal ions on the growth and DAE expression. The results indicated that the optimal medium formulation as: sucrose 10 g/L, soybean peptone 15 g/L,(NH4)2SO4 3 g/L, KH2PO4 3 g/L, MgSO4 0.5 g/L, MnSO4 0.025 mmol/L. On this basis, the cultivation conditions were further studied through single factor experiments. The optimal cultivation conditions were liquid loading amount of 30% in a flask, inoculation volume of 3%, IPTG concentration of 1 mmol/L, and induction time of 10 h. Under this condition, the OD600 of recombinant E. coli had a 1.46-fold increase, and the volume DAE activity had a 2.57-fold increase compared to the starting culture conditions. The culture medium and condition were further tested by batch fermentation in a 5 L bioreactor. After 18 h of induction, the OD600 of recombinant cells reached the highest value of 51.8, the dry cell weight reached 21.5 g/L, and the volume enzyme activity reached 103.8 U/mL. The reactions from 500 g/L D-fructose to D-allulose were carried out with different recombinant cell concentrations at pH 7, 50℃. When the cell concentration was 0.014 g DCW/L, the product D-allulose reached the highest concentration at 149.74 g/L and conversion rate at 28.76% after 1 h reaction. This study has reference value for the fermentative production of DAE.
LIU Weiwei
,
CHANG Chuting
,
FENG Jingjie
,
ZHANG Jiahe
,
LI Feisheng
,
DING Wentao
,
WANG Changlu
. Optimization of fermentation conditions of recombinant Escherichia coli to improve the production of D-allulose 3-epimerase[J]. Food and Fermentation Industries, 2024
, 50(4)
: 253
-259
.
DOI: 10.13995/j.cnki.11-1802/ts.035843
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