Plug-in repressor library for precise regulation of metabolic flux in Escherichia coli |
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| Institution: | 1. Department of Chemical and Biomolecular Engineering, The University of Tennessee, Knoxville, TN, United States;2. Center for Bioenergy Innovation, Oak Ridge National Laboratory, Oak Ridge, TN, United States;1. Department of Chemical Engineering and Pohang University of Science and Technology, 77 Cheongam-Ro, Nam-Gu, Pohang, Gyeongbuk 37673, Republic of Korea;2. School of Interdisciplinary Bioscience and Bioengineering, Pohang University of Science and Technology, 77 Cheongam-Ro, Nam-Gu, Pohang, Gyeongbuk 37673, Republic of Korea;3. School of Energy and Chemical Engineering, Ulsan National Institute of Science and Technology, 50, UNIST-gil, Ulsan 44919, Republic of Korea;4. School of Chemical and Biological Engineering, Institute of Chemical Process, Seoul National University, 1 Gwanak-Ro, Gwanak-Gu, Seoul 08826, Republic of Korea;1. Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodai, Nada, Kobe, 657-8501, Japan;2. Department of Biomolecular Engineering Graduate School of Engineering, Tohoku University, 6-6-11 Aoba, Aramaki, Aoba-ku, Sendai, 980-8579, Japan;3. Center for Sustainable Resource Science, RIKEN, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa, 230-0045, Japan;4. Graduate School of Science, Technology and Innovation, Kobe University, 1-1 Rokkodai, Nada, Kobe, 657-8501, Japan |
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| Abstract: | In metabolic engineering, enhanced production of value-added chemicals requires precise flux control between growth-essential competing and production pathways. Although advances in synthetic biology have facilitated the exploitation of a number of genetic elements for precise flux control, their use requires expensive inducers, or more importantly, needs complex and time-consuming processes to design and optimize appropriate regulator components, case-by-case. To overcome this issue, we devised the plug-in repressor libraries for target-specific flux control, in which expression levels of the repressors were diversified using degenerate 5′ untranslated region (5’ UTR) sequences employing the UTR Library Designer. After we validated a wide expression range of the repressor libraries, they were applied to improve the production of lycopene from glucose and 3-hydroxypropionic acid (3-HP) from acetate in Escherichia coli via precise flux rebalancing to enlarge precursor pools. Consequently, we successfully achieved optimal carbon fluxes around the precursor nodes for efficient production. The most optimized strains were observed to produce 2.59 g/L of 3-HP and 11.66 mg/L of lycopene, which were improved 16.5-fold and 2.82-fold, respectively, compared to those produced by the parental strains. These results indicate that carbon flux rebalancing using the plug-in library is a powerful strategy for efficient production of value-added chemicals in E. coli. |
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| Keywords: | Metabolic engineering Carbon flux rebalancing 3-Hydroxypropionic acid Lycopene UTR Library designer |
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