A novel enantioselective epoxide hydrolase from Agromyces mediolanus ZJB120203: Cloning,characterization and application |
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| Institution: | 1. Institute of Bioengineering, Zhejiang University of Technology, Hangzhou 310014, Zhejiang, PR China;2. Engineering Research Center of Bioconversion and Biopurification of the Ministry of Education, Zhejiang University of Technology, Hangzhou 310014, Zhejiang, PR China;1. College of Life Science, Department of Materials Chemistry, Huzhou University, Huzhou 313000, PR China;2. Institute of Bioengineering, College of Chemical and Biological Engineering, Zhejiang University, Hangzhou 310027, PR China;1. Environmental Biotechnology and Genomics Division, CSIR-NEERI, Nagpur, India;2. Environmental Material Division, CSIR-NEERI, Nagpur, India;3. Solid and Hazardous Waste Management Division, CSIR-NEERI, Nagpur, India;4. Academy of Scientific and Innovative Research, CSIR-NEERI, Nagpur, 440020 Maharashtra, India;1. Key Laboratory of Environmental and Applied Microbiology, Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu 610041, China;2. Environmental Microbiology Key Laboratory of Sichuan Province, Chengdu 610041, China;3. University of the Chinese Academy of Sciences, Beijing 100049, China;1. Department of Biochemistry, Groningen Biomolecular Science and Biotechnology Institute, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands;2. School of Life Science and Technology, University of Electronic Science and Technology of China, Chengdu, China;3. Ru?er Bo?kovi? Institute, Bijeni?ka c. 54, 10000 Zagreb, Croatia;4. BASF SE, Biological & Effect Systems Research, Ludwigshafen, Germany |
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| Abstract: | A new strain Agromyces mediolanus ZJB120203, capable of enantioselective epoxide hydrolase (EH) activity was isolated employing a newly established colorimetric screening and chiral GC analysis method. The partial nucleotide sequence of an epoxide hydrolase (AmEH) gene from A. mediolanus ZJB120203 was obtained by PCR using degenerate primers designed based on the conserved domains of EHs. Subsequently, an open reading frame containing 1167 bp and encoding 388 amino acids polypeptide were identified. Expression of AmEH was carried out in Escherichia coli and purification was performed by Nickel-affinity chromatography. The purified AmEH had a molecular weight of 43 kDa and showed its optimum pH and temperature at 8.0 and 35 °C, respectively. Moreover, this AmEH showed broad substrates specificity toward epoxides. In this study, it is demonstrated that the AmEH could unusually catalyze the hydrolysis of (R)-ECH to produce enantiopure (S)-ECH. Enantiopure (S)-ECH could be obtained with enantiomeric excess (ee) of >99% and yield of 21.5% from 64 mM (R,S)-ECH. It is indicated that AmEH from A. mediolanus is an attractive biocatalyst for the efficient preparation of optically active ECH. |
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| Keywords: | Cloning Characterization Epoxide hydrolase |
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