Niu Panqing, Dong Xiaoxiang, Wang Yuancai, Liu Liming
State Key Laboratory of Food Science and Technology, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; Laboratory of Food Microbial-Manufacturing Engineering, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China.
State Key Laboratory of Food Science and Technology, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; Laboratory of Food Microbial-Manufacturing Engineering, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China.
J Biotechnol. 2014 Jun 10;179:56-62. doi: 10.1016/j.jbiotec.2014.03.021. Epub 2014 Mar 21.
In this study, a novel strategy for α-ketoglutaric acid (α-KG) production from l-glutamic acid using recombinant l-glutamate oxidase (LGOX) was developed. First, by analyzing the molecular structure characteristics of l-glutamic acid and α-KG, LGOX was found to be the best catalyst for oxidizing the amino group of l-glutamic acid to a ketonic group without the need for exogenous cofactor. Then the LGOX gene was expressed in Escherichia coli BL21 (DE3) in a soluble and active form, and the recombinant LGOX activity reached to a maximum value of 0.59U/mL at pH 6.5, 30°C. Finally, the maximum α-KG concentration reached 104.7g/L from 110g/L l-glutamic acid in 24h, under the following optimum conditions: 1.5U/mL LGOX, 250U/mL catalase, 3mM MnCl2, 30°C, and pH 6.5.
在本研究中，开发了一种利用重组L-谷氨酸氧化酶（LGOX）从L-谷氨酸生产α-酮戊二酸（α-KG）的新策略。首先，通过分析L-谷氨酸和α-KG的分子结构特征，发现LGOX是将L-谷氨酸的氨基氧化为酮基的最佳催化剂，无需外源辅因子。然后，LGOX基因在大肠杆菌BL21（DE3）中以可溶且有活性的形式表达，重组LGOX活性在pH 6.5、30°C时达到最大值0.59U/mL。最后，在以下最佳条件下：1.5U/mL LGOX、250U/mL过氧化氢酶、3mM MnCl2、30°C和pH 6.5，24小时内从110g/L L-谷氨酸中获得的最大α-KG浓度达到104.7g/L。
