Production of glucaric acid from myo-inositol in engineered Pichia pastoris |
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| Affiliation: | 1. The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, China;2. Synergetic Innovation Center of Food Safety and Nutrition, 1800 Lihu Road, Wuxi, Jiangsu 214122, China;3. School of Biotechnology, Jiangnan University, Wuxi 214122, China;1. Beijing Bioprocess Key Laboratory, State Key Laboratory of Chemical Resource Engineering, Beijing University of Chemical Technology, Beijing 100029, PR China;2. Amoy - BUCT Industrial Bio-technovation Institute, Amoy 361022, PR China;1. State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, P.O. Box 329, 130 Meilong Road, Shanghai 200237, China;2. Shan Dong Fuyang Biological Technology Co., Ltd, China;1. Key Laboratory of Biomass Chemical Engineering of Ministry of Education, Department of Chemical and Biological Engineering, Zhejiang University, Hangzhou 310027, China;2. Center for Systems Biology, University of Iceland, Reykjavik, Iceland;1. National Engineering Laboratory for Cereal Fermentation Technology (NELCF), Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China;2. The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China;3. School of Pharmaceutical Science, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China;1. Jiangxi Provincial Key Laboratory of Molecular Medicine, The Second Affiliated Hospital of Nanchang University, Nanchang, PR China;2. School of Pharmaceutical Science, Nanchang University, Nanchang 330006, PR China;3. State Key Laboratory of Food Science and Technology, Institute for Advanced Study, University of Nanchang, Nanchang 330047, PR China |
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| Abstract: | A potential myo-inositol oxygenase (ppMIOX) was identified as a functional enzyme and a glucaric acid synthetic pathway was firstly constructed in Pichia pastoris. Coexpression of the native ppMIOX and the urinate dehydrogenase (Udh) from Pseudomonas putida KT2440 led to obvious accumulation of glucaric acid (90.46 ± 0.04 mg/L) from myo-inositol whereas no glucaric acid was detected from glucose. In comparison, coexpression of the heterologous mouse MIOX (mMIOX) and Udh resulted in higher titers of glucaric acid from glucose and myo-inositol, 107.19 ± 11.91 mg/L and 785.4 ± 1.41 mg/L, respectively. By applying a fusion expression strategy with flexible peptides, the mMIOX specific activity and the glucaric acid concentration were significantly increased. Using glucose and myo-inositol as carbon substrates, the production of glucaric acid was substantially enhanced to 6.61 ± 0.30 g/L in fed-batch cultures. To the best of our knowledge, this is the highest reported value to date. |
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| Keywords: | Glucaric acid MIOX Metabolic engineering |
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