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过程工程学报 ›› 2016, Vol. 16 ›› Issue (2): 282-297.DOI: 10.12034/j.issn.1009-606X.215338

• 生化工程专栏 • 上一篇    下一篇

L-谷氨酸氧化酶高密度发酵及催化合成α-酮戊二酸

樊祥臣 陈瑞东 刘佳 徐继嗣 刘立明   

  1. 江南大学生物工程学院食品科学与技术国家重点实验室 江南大学生物工程学院食品科学与技术国家重点实验室 江南大学生物工程学院食品科学与技术国家重点实验室 江南大学生物学院食品科学与技术国家重点实验室 江南大学生物工程学院环境生物技术实验室
  • 收稿日期:2015-10-10 修回日期:2015-11-30 出版日期:2016-04-20 发布日期:2016-12-22
  • 通讯作者: 樊祥臣

High Cell Density Fermentation of L-Glutamate Oxidase and Its Catalysis for Synthesis of a-Ketoglutaric Acid

FAN Xiang-chen CHEN Rui-dong LIU Jia XU Ji-si LI Zhen   

  1. State Key Laboratory of Food Science and Technology, Jiangnan Universit State Key Laboratory of Food Science and Technology, Jiangnan Universit State Key Laboratory of Food Science and Technology, Jiangnan Universit State Key Laboratory of Food Science and Technology, Jiangnan Universit Lab of Biotechnology, School of Biotechnology, Southern Yangtze University
  • Received:2015-10-10 Revised:2015-11-30 Online:2016-04-20 Published:2016-12-22
  • Contact: FAN Xiang-chen

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摘要： 以乳糖代替异丙基硫代半乳糖苷(IPTG)作为诱导剂，优化补料策略提高菌体密度，优化诱导策略提高蛋白表达，以高密度发酵所得L-谷氨酸氧化酶(LGOX)全细胞转化L-谷氨酸生产a-酮戊二酸(a-KG). 结果表明，摇瓶培养条件下LGOX酶活由IPTG诱导的3.12 U/mL提高到4.78 U/mL；通过指数补料与DO-stat补料相结合的两阶段补料策略，5 L发酵罐中细胞浓度由IPTG诱导的2.49 g/L提高到41.6 g/L, LGOX酶活为59 U/mL. 优化诱导策略后细胞浓度为48.4 g/L, LGOX酶活提高到156.1 U/mL，分别是优化前的19.4和50倍，a-KG产量达5160 g/L，提高48倍.

关键词: L-谷氨酸氧化酶, a-酮戊二酸, 发酵, 全细胞转化

Abstract: Lactose was used to replace isopropyl β-D-1-thiogalactopyranoside (IPTG) as inducer, the feeding strategy was optimized to improve cell density, induction strategy was studied for improvement of L-glutamate oxidase (LGOX) activity, the whole-cell conversion biocatalyst from high cell density fermentation was used to produce a-ketoglutaric acid (a-KG) from L-glutamic acid. The results showed that LGOX activity was up to 4.78 U/mL from 3.12 U/mL by IPTG induction in the shake flask. In a 5 L fermenter, cell density was increased from 2.49 to 41.6 g/L, and LGOX activity reached 59 U/mL with two-stage feeding strategy combining the exponential feeding with DO-stat feeding strategy. By optimizing the induction strategies, the maximal cell density and LGOX activity reached 48.4 g/L and 156.1 U/mL, improvement of 19.4- and 50-fold before optimization, respectively. a-KG yield was up to 5160 g/L by whole-cell conversion, increased by 48-fold.

Key words: L-glutamate oxidase, a-ketoglutaric acid, fermentation, whole-cell conversion

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