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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?-酮戊二酸(?-KG).结果表明,摇瓶培养条件下LGOX酶活由IPTG诱导的3.12 U/m L提高到4.78 U/m L;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/m L.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/m L,分别是优化前的19.4和50倍,?-KG产量达5160 g/L,提高48倍.
Lactose was used instead of isopropylthiogalactoside (IPTG) as an inducing agent to optimize the feeding strategy to increase the cell density and optimize the induction strategy to improve the protein expression. The L-glutamate oxidase (LGOX) obtained by high-density fermentation L-glutamic acid was used to produce? -ketoglutarate (? -KG). The results showed that LGOX enzyme activity increased from 3.12 U / m L induced by IPTG to 4.78 U / m L in shake flask culture, In the two-stage feeding strategy combining feed and DO-stat feed, the cell concentration in 5 L fermenter was increased from 2.49 g / L to 41.6 g / L induced by IPTG and the activity of LGOX was 59 U / m L. Optimization After induction, the cell concentration was 48.4 g / L and the activity of LGOX enzyme increased to 156.1 U / mL, which was 19.4 and 50 times before optimization, and the yield of? KG was up to 4800.