Improving metabolic efficiency of the reverse beta-oxidation cycle by balancing redox cofactor requirement |
| |
| Institution: | 1. College of Food Science and Technology, Nanjing Agricultural University, 1 Weigang Road, Nanjing, Jiangsu 210095, China;2. Institute of Agro-Product Processing, Jiangsu Academy of Agricultural Sciences, Nanjing, Jiangsu 210095, China;1. Department of Genetic Engineering, College of Biotechnology and Bioengineering, Sungkyunkwan University, Suwon 440-746, Republic of Korea;2. School of Interdisciplinary Bioscience and Bioengineering, Pohang University of Science and Technology, Pohang 790-784, Republic of Korea;3. Department of Chemical Engineering, Pohang University of Science and Technology, Pohang 790-784, Republic of Korea;4. Department of Agricultural Biotechnology and Center for Agricultural Biomaterials, Seoul National University, Seoul 151-921, Republic of Korea;5. Bio R&D Center, Samsung Advanced Institute of Technology, Yongin 446-712, Republic of Korea;6. Department of Food Science and Engineering, Ewha Womans University, Seoul 120-750, Republic of Korea;7. Department of Bioengineering and Technology, College of Engineering, Kangwon National University, Chuncheon 220-701, Republic of Korea;1. Department of Chemical and Biomolecular Engineering, Rice University, Houston, TX, USA;2. Department of Bioengineering, Rice University, Houston, TX, USA;1. School of Life Sciences, Ulsan National Institute of Science and Technology (UNIST), Ulsan 44919, Republic of Korea;2. School of Energy and Chemical Engineering, Ulsan National Institute of Science and Technology (UNIST), Ulsan 44919, Republic of Korea;1. Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, WI, United States;2. Microbiology Doctoral Training Program, University of Wisconsin-Madison, Madison, WI, United States;3. Department of Biology and Biological Engineering, Chalmers University of Technology, Göteborg, Sweden |
| |
| Abstract: | Previous studies have made many exciting achievements on pushing the functional reversal of beta-oxidation cycle (r-BOX) to more widespread adoption for synthesis of a wide variety of fuels and chemicals. However, the redox cofactor requirement for the efficient operation of r-BOX remains unclear. In this work, the metabolic efficiency of r-BOX for medium-chain fatty acid (C6-C10, MCFA) production was optimized by redox cofactor engineering. Stoichiometric analysis of the r-BOX pathway and further experimental examination identified NADH as a crucial determinant of r-BOX process yield. Furthermore, the introduction of formate dehydrogenase from Candida boidinii using fermentative inhibitor byproduct formate as a redox NADH sink improved MCFA titer from initial 1.2 g/L to 3.1 g/L. Moreover, coupling of increasing the supply of acetyl-CoA with NADH to achieve fermentative redox balance enabled product synthesis at maximum titers. To this end, the acetate re-assimilation pathway was further optimized to increase acetyl-CoA availability associated with the new supply of NADH. It was found that the acetyl-CoA synthetase activity and intracellular ATP levels constrained the activity of acetate re-assimilation pathway, and 4.7 g/L of MCFA titer was finally achieved after alleviating these two limiting factors. To the best of our knowledge, this represented the highest titer reported to date. These results demonstrated that the key constraint of r-BOX was redox imbalance and redox engineering could further unleash the lipogenic potential of this cycle. The redox engineering strategies could be applied to acetyl-CoA-derived products or other bio-products requiring multiple redox cofactors for biosynthesis. |
| |
| Keywords: | β-oxidation reversal ATP NADH Synthetic biology Metabolic engineering |
| 本文献已被 ScienceDirect 等数据库收录！ |
|
