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Systematic engineering of TCA cycle for optimal production of a four-carbon platform chemical 4-hydroxybutyric acid in Escherichia coli
Affiliation:1. Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus Program), Center for Systems and Synthetic Biotechnology, Institute for the BioCentury, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 34141, Republic of Korea;2. BioInformatics Research Center, KAIST, Daejeon 34141, Republic of Korea;3. BioProcess Engineering Research Center, KAIST, Daejeon 34141, Republic of Korea;1. Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, NC 27695-7905, United States;2. Dept. of Biochemistry and Molecular Biology, University of Georgia, Athens, GA 30602, United States;1. Beijing Key Laboratory of Bioprocess, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing 100029, China;2. Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, United States;3. School of Life Sciences, Tsinghua University, Beijing 100084, China;1. Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus program), BioProcess Engineering Research Center, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea;2. Center for Systems and Synthetic Biotechnology, Institute for the BioCentury, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea;1. State Key Laboratory of Chemical Resource Engineering, Beijing University of Chemical Technology, Beijing 100029, China;2. Beijing Advanced Innovation Center for Soft Matter Science and Engineering, Beijing University of Chemical Technology, Beijing 100029, China;3. College of Engineering, The University of Georgia, Athens, GA 30602, USA;4. Department of Chemical and Biomolecular Engineering, University of California, Los Angeles, 420 Westwood Plaza, Los Angeles, CA 90095, USA;5. UCLA-DOE Institute of Genomics and Proteomics, 420 Westwood Plaza, Los Angeles, CA 90095, USA;1. Institute of Applied Chemistry, Department of Chemical Engineering, Tsinghua University, Beijing 100084, China;2. Tsinghua Innovation Center in Dongguan, Dongguan 523808, China
Abstract:To address climate change and environmental problems, it is becoming increasingly important to establish biorefineries for the production of chemicals from renewable non-food biomass. Here we report the development of Escherichia coli strains capable of overproducing a four-carbon platform chemical 4-hybroxybutyric acid (4-HB). Because 4-HB production is significantly affected by aeration level, genome-scale metabolic model-based engineering strategies were designed under aerobic and microaerobic conditions with emphasis on oxidative/reductive TCA branches and glyoxylate shunt. Several different metabolic engineering strategies were employed to develop strains suitable for fermentation both under aerobic and microaerobic conditions. It was found that microaerobic condition was more efficient than aerobic condition in achieving higher titer and productivity of 4-HB. The final engineered strain produced 103.4 g/L of 4-HB by microaerobic fed-batch fermentation using glycerol. The aeration-dependent optimization strategy of TCA cycle will be useful for developing microbial strains producing other reduced derivative chemicals of TCA cycle intermediates.
Keywords:Systems metabolic engineering  4-hydroxybutyric acid  TCA cycle engineering  Microaerobic condition  Metabolic simulation
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