Engineering improved ethanol production in Escherichia coli with a genome-wide approach |
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| Institution: | 1. Global Technology Center, Samsung Electronics Co., Ltd., Suwon, Gyeonggi-do 443-742, Republic of Korea;2. Light Metal Division, Korea Institute of Materials Science, 797 Changwondaero, Seongsan-gu, Changwon, Gyeongnam 642-831, Republic of Korea;3. IFW Dresden, Institute for Complex Materials, P.O. Box 27 01 16, D-01171 Dresden, Germany;4. TU Dresden, Institute of Materials Science, D-01062 Dresden, Germany;5. Center for Non-crystalline Materials, Department of Materials Science and Engineering, Yonsei University, 134 Shinchon-dong, Seodaemun-gu, Seoul 120-749, Republic of Korea;1. Department of Chemical Engineering, University of Waterloo, 200 University Avenue West, Waterloo, Ontario N2L3G1, Canada;2. Waterloo Institute for Nanotechnology, University of Waterloo, 200 University Avenue West, Waterloo, Ontario N2L3G1, Canada;3. Institute of Optoelectronic Technology, Beijing Jiaotong University, Number 3, Shangyuancun, Haidian District, Beijing 100044, People’s Republic of China;1. Department of Urology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China;2. Institute of Urology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China;3. Department of Pathology and Urology, Jonsson Comprehensive Cancer Center and Broad Center of Regenerative Medicine and Stem Cell Research, UCLA David Geffen School of Medicine, Los Angeles, CA |
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| Abstract: | A key challenge to the commercial production of commodity chemical and fuels is the toxicity of such molecules to the microbial host. While a number of studies have attempted to engineer improved tolerance for such compounds, the majority of these studies have been performed in wild-type strains and culturing conditions that differ considerably from production conditions. Here we applied the multiscalar analysis of library enrichments (SCALEs) method and performed a growth selection in an ethanol production system to quantitatively map in parallel all genes in the genome onto ethanol tolerance and production. In order to perform the selection in an ethanol-producing system, we used a previously engineered Escherichia coli ethanol production strain (LW06; ATCC BAA-2466) (Woodruff et al., in press), as the host strain for the multiscalar genomic library analysis (>106 clones for each library of 1, 2, or 4 kb overlapping genomic fragments). By testing individually selected clones, we confirmed that growth selections enriched for clones with both improved ethanol tolerance and production phenotypes. We performed combinatorial testing of the top genes identified (uspC, otsA, otsB) to investigate their ability to confer improved ethanol tolerance or ethanol production. We determined that overexpression of otsA was required for improved tolerance and productivity phenotypes, with the best performing strains showing up to 75% improvement relative to the parent production strain. |
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