Abstract
Microbial-produced branched-chain higher alcohols (BCHAs), such as isopropanol, isobutanol, and isopentanol in Escherichia coli, have emerged as promising alternative biofuels under development. Elucidating and improving the tolerance of E. coli to BCHAs are important issues for microbial production of BCHAs due to their physiological inhibitory effect. Previous works aimed at understanding the genetic basis of E. coli tolerance to BCHAs with a comparative genome, reverse engineering, or transcriptome approach have gained some important insights into the mechanism of tolerance. However, investigation on BCHA tolerance from the whole-genomic, transcriptomic, and metabolic levels via a systematic approach has not yet been completely elucidated. Here, in this study, genomic, transcriptomic, and 13C-metabolic flux analyses (13C-MFA) of an evolved E. coli strain adapted to BCHA tolerance were conducted. Genome mutation of negative regulation factor (rssB, acrB, and clpX) of RpoS level suggested upregulation of RpoS activity in BCHA tolerance of E. coli. From a more detailed perspective, enhanced energy metabolism was observed to be the main characteristic of E. coli strain tolerant to BCHAs. Enhanced energy metabolism has been achieved through several routes, which included redistribution of the central carbon metabolism, upregulation of the energy generation machinery, and facilitating the operation of electron transferring chain. Evidence of multiple solutions of genotype modification toward BCHA tolerance was also revealed through comparative analysis of previous works from different groups.
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Acknowledgments
We would like to thank Dr. Le You and Dr. Lian He from the Washington University in St. Louis for discussion on 13C-metabolic flux analysis experiments. We also would like to thank Ms. Xianni Qi and Yuanyuan Zhang of Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, for technique assistance in GS-MS analysis.
Funding
This work was supported by the National Basic Research Program (973 Program, 2011CBA00800) and the National Knowledge Innovation Project of the Chinese Academy of Sciences (KSCX2-EW-G-14). QHW was supported by the One Hundred Talent Program of the Chinese Academy of Sciences, and RT was supported by Youth Innovation Promotion Association of Chinese Academy of Sciences.
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Wang, B., Guo, Y., Xu, Z. et al. Genomic, transcriptomic, and metabolic characterizations of Escherichia coli adapted to branched-chain higher alcohol tolerance. Appl Microbiol Biotechnol 104, 4171–4184 (2020). https://doi.org/10.1007/s00253-020-10507-0
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DOI: https://doi.org/10.1007/s00253-020-10507-0