Zhu Xiaoyan, Gong Jinsong, Li Heng, Lu Zhenming, Zhou Zhemin, Shi Jinsong, Xu Zhenghong
Sheng Wu Gong Cheng Xue Bao. 2014 Mar;30(3):412-24.
Microbial nitrilases have attracted increasing attention in nitrile hydrolysis for carboxylic acid production in recent years. A bacterium with nitrilase activity was isolated and identified as Pseudomonas putida CGMCC3830 based on its morphology, physiological and biochemical characteristics, as well as 16S rRNA gene sequence. The nitrilase production was optimized by varying culture conditions using the one-factor-at-a-time method and response surface methodology. Glycerol 13.54 g/L, tryptone 11.59 g/L, yeast extract 5.21 g/L, KH2PO4 1 g/L, NaCl 1 g/L, urea 1 g/L, initial pH 6.0 and culture temperature 30 degrees C were proved to be the optimal culture conditions. It resulted in the maximal nitrilase production of 36.12 U/mL from 2.02 U/mL. Investigations on substrate specificity demonstrate P. putida nitrilase preferentially hydrolyze aromatic nitriles. When applied in nicotinic acid synthesis, 2 mg/mL P. putida cells completely hydrolyzed 20.8 g/L 3-cyanopyridine into nicotinic acid in 90 min. The results indicated P. putida CGMCC3830 displayed potential for industrial production of nicotinic acid.
近年来，微生物腈水解酶在用于生产羧酸的腈水解反应中受到越来越多的关注。基于其形态、生理生化特性以及16S rRNA基因序列，分离出一株具有腈水解酶活性的细菌，并鉴定为恶臭假单胞菌CGMCC3830。采用单因素法和响应面法通过改变培养条件对腈水解酶的生产进行了优化。结果表明，甘油13.54 g/L、胰蛋白胨11.59 g/L、酵母提取物5.21 g/L、KH2PO4 1 g/L、NaCl 1 g/L、尿素1 g/L、初始pH 6.0和培养温度30℃为最佳培养条件。在此条件下，腈水解酶产量从2.02 U/mL提高到36.12 U/mL。对底物特异性的研究表明，恶臭假单胞菌腈水解酶优先水解芳香腈。在用于烟酸合成时，2 mg/mL的恶臭假单胞菌细胞在90分钟内可将20.8 g/L的3-氰基吡啶完全水解为烟酸。结果表明，恶臭假单胞菌CGMCC3830在烟酸的工业化生产中具有潜力。
