大肠杆菌ptsG基因缺陷菌株的构建及其发酵混合糖产L-丙氨酸

为了增强大肠杆菌（Escherichia coli）JH-B2利用混合糖产L-丙氨酸的能力，通过Red同源重组技术敲除了转运葡萄糖的关键基因ptsG，构建新菌株JH-B3。结果表明，以10%混合糖（5%葡萄糖和5%木糖）为碳源，ptsG基因缺陷菌株JH-B3同时利用葡萄糖和木糖，且在32 h时利用完葡萄糖，表明了ptsG基因的敲除大幅减弱了葡萄糖效应；而菌株JH-B2在发酵24 h时利用完葡萄糖，在28 h时才开始利用木糖，表现出葡萄糖效应。发酵至98 h，ptsG基因缺陷菌株JH-B3已经利用完木糖，其丙氨酸产量为94.1 g/L，生产强度为0.96 g/（L·h），而菌株JH-B2在同等时间内还剩18.10 g/L木糖未利用，其丙氨酸产量为77.6 g/L，生产强度为0.79 g/（L·h）。ptsG基因缺陷菌株JH-B3的丙氨酸生产强度较JH-B2提高了21.5%，能够同时利用混合糖发酵产丙氨酸，为利用木质纤维素糖源生产丙氨酸提供了工业应用基础。

Construction of ptsG gene-deficient strain of Escherichia coli and L-alanine production by mixed sugars fermentation

In order to enhance the L-alanine-producing ability of Escherichia coli JH-B2 by using mixed sugar, the new strain JH-B3 was constructed by knockout ptsG (a key glucose transporter gene) using Red homologous recombination technique. The results showed that using 10% mixed sugars (glucose 5% and xylose 5%) as carbon source, the ptsG gene-deficient strain JH-B3 could simultaneously use glucose and xylose, and the glucose was used up at 32 h, which indicated that the glucose effect was greatly weakened due to the knockout of ptsG gene. However, the strain JH-B2 used up glucose at 24 h, and started to use xylose at 28 h, showing the glucose effect. Furthermore, the ptsG gene-deficient strain JH-B3 had used up xylose at 98 h, and the alanine yield and productivity were 94.1 g/L and 0.96 g/(L·h), respectively. In comparison, the strain JH-B2 was unable to use up xylose at 98 h (with 18.10 g/L of xylose remaining), the alanine yield and productivity were 77.6 g/L and 0.79 g/(L·h), respectively. The alanine productivity of the ptsG gene-deficient strain JH-B3 was 21.5% higher than that of strain JH-B2, and the ptsG gene-deficient strain JH-B3 was able to produce alanine by simultaneously using mixed sugars, which provided an industrial application basis for the production of alanine by lignocellulose sugar source.