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Hu B, Yang YM, Beck DA, et al. Comprehensive molecular characterization of Methylobacterium extorquens AM1 adapted for 1-butanol tolerance. Biotechnol Biofuels. 2016;9:84doi: 10.1186/s13068-016-0497-y.
Hu, B., Yang, Y. M., Beck, D. A., Wang, Q. W., Chen, W. J., Yang, J., Lidstrom, M. E., & Yang, S. (2016). Comprehensive molecular characterization of Methylobacterium extorquens AM1 adapted for 1-butanol tolerance. Biotechnology for biofuels, 984. https://doi.org/10.1186/s13068-016-0497-y
Hu, Bo, et al. "Comprehensive molecular characterization of Methylobacterium extorquens AM1 adapted for 1-butanol tolerance." Biotechnology for biofuels vol. 9 (2016): 84. doi: https://doi.org/10.1186/s13068-016-0497-y
Hu B, Yang YM, Beck DA, Wang QW, Chen WJ, Yang J, Lidstrom ME, Yang S. Comprehensive molecular characterization of Methylobacterium extorquens AM1 adapted for 1-butanol tolerance. Biotechnol Biofuels. 2016 Apr 11;9:84. doi: 10.1186/s13068-016-0497-y. eCollection 2016. PMID: 27069508; PMCID: PMC4827201.
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Biotechnol Biofuels. 2016 Apr 11;9:84. doi: 10.1186/s13068-016-0497-y. eCollection 2016.

Comprehensive molecular characterization of Methylobacterium extorquens AM1 adapted for 1-butanol tolerance.

Biotechnology for biofuels

Bo Hu, Yi-Ming Yang, David A C Beck, Qian-Wen Wang, Wen-Jing Chen, Jing Yang, Mary E Lidstrom, Song Yang

Affiliations

  1. Department of Chemical Engineering, University of Washington, Seattle, WA USA ; Industrial Product Division, Intrexon Corporation, South San Francisco, CA 94080 USA.
  2. School of Life Science, Shandong Province Key Laboratory of Applied Mycology, and Qingdao International Center on Microbes Utilizing Biogas, Qingdao Agricultural University, Qingdao, Shandong Province China.
  3. Department of Chemical Engineering, University of Washington, Seattle, WA USA ; eScience Institute, University of Washington, Seattle, WA USA.
  4. Central Laboratory, Qingdao Agricultural University, Qingdao, Shandong Province China.
  5. Department of Chemical Engineering, University of Washington, Seattle, WA USA ; Department of Microbiology, University of Washington, Seattle, WA 98195-1750 USA.
  6. School of Life Science, Shandong Province Key Laboratory of Applied Mycology, and Qingdao International Center on Microbes Utilizing Biogas, Qingdao Agricultural University, Qingdao, Shandong Province China ; Key Laboratory of Systems Bioengineering, Ministry of Education, Tianjin University, Tianjin, China.

PMID: 27069508 PMCID: PMC4827201 DOI: 10.1186/s13068-016-0497-y

Abstract

BACKGROUND: The toxicity of alcohols is one of the major roadblocks of biological fermentation for biofuels production. Methylobacterium extorquens AM1, a facultative methylotrophic α-proteobacterium, has been engineered to generate 1-butanol from cheap carbon feedstocks through a synthetic metabolic pathway. However, M. extorquens AM1 is vulnerable to solvent stress, which impedes further development for 1-butanol production. Only a few studies have reported the general stress response of M. extorquens AM1 to solvent stress. Therefore, it is highly desirable to obtain a strain with ameliorated 1-butanol tolerance and elucidate the molecular mechanism of 1-butnaol tolerance in M. extorquens AM1 for future strain improvement.

RESULTS: In this work, adaptive laboratory evolution was used as a tool to isolate mutants with 1-butanol tolerance up to 0.5 %. The evolved strains, BHBT3 and BHBT5, demonstrated increased growth rates and higher survival rates with the existence of 1-butanol. Whole genome sequencing revealed a SNP mutation at kefB in BHBT5, which was confirmed to be responsible for increasing 1-butanol tolerance through an allelic exchange experiment. Global metabolomic analysis further discovered that the pools of multiple key metabolites, including fatty acids, amino acids, and disaccharides, were increased in BHBT5 in response to 1-butanol stress. Additionally, the carotenoid synthesis pathway was significantly down-regulated in BHBT5.

CONCLUSIONS: We successfully screened mutants resistant to 1-butanol and provided insights into the molecular mechanism of 1-butanol tolerance in M. extorquens AM1. This research will be useful for uncovering the mechanism of cellular response of M. extorquens AM1 to solvent stress, and will provide the genetic blueprint for the rational design of a strain of M. extorquens AM1 with increased 1-butanol tolerance in the future.

Keywords: 1-Butanol tolerance; Adaptive evolution; Carotenoid; Global metabolome analysis; Methylobacterium extorquens AM1; Whole genome sequencing

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