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Applied Microbiology and Biotechnology

, Volume 103, Issue 1, pp 239–250 | Cite as

Engineering and manipulation of a mevalonate pathway in Escherichia coli for isoprene production

  • Chun-Li Liu
  • Hao-Ran Bi
  • Zhonghu Bai
  • Li-Hai Fan
  • Tian-Wei Tan
Biotechnological products and process engineering

Abstract

Isoprene is a useful phytochemical with high commercial values in many industrial applications including synthetic rubber, elastomers, isoprenoid medicines, and fossil fuel. Currently, isoprene is on large scale produced from petrochemical sources. An efficient biological process for isoprene production utilizing renewable feedstocks would be an important direction of research due to the fossil raw material depletion and air pollution. In this study, we introduced the mevalonate (MVA) pathway genes/acetoacetyl-coenzyme A thiolase (mvaE) and MVA synthase (mvaS) from Enterococcus faecalis (E. faecalis); MVA kinase (mvk) derived from Methanosarcina mazei (M. mazei); and phosphomevalonate kinase (pmk), diphosphomevalonate decarboxylase (mvaD), and isopentenyl diphosphate isomerase (idi) from Streptococcus pneumoniae (S. pneumoniae) to accelerate dimethylallyl diphosphate (DMAPP) accumulation in Escherichia coli (E. coli). Together with a codon-optimized isoprene synthase (ispS) from Populus alba (P. alba), E. coli strain succeeded in formation of isoprene. We then manipulated the heterologous MVA pathway for high-level production of isoprene, by controlling the gene expression levels of the MVA pathway genes. We engineered four E. coli strains which showed different gene expression levels and different isoprene productivities, and we also characterized them with quantitative real-time PCR and metabolite analysis. To further improve the isoprene titers and release the toxicity to cells, we developed the extraction fermentation by adding dodecane in cultures. Finally, strain BL2T7P1TrcP harboring balanced gene expression system produced 587 ± 47 mg/L isoprene, with a 5.2-fold titer improvement in comparison with strain BL7CT7P. This work indicated that a balanced metabolic flux played a significant role to improve the isoprene production via MVA pathway.

Keywords

Isoprene Mevalonate pathway Balanced gene expression Escherichia coli 

Notes

Acknowledgments

We thank Xiaozhou Luo (Lawrence Berkeley National Lab, Joint Bio-Energy Institute) for helping to revise this manuscript.

Authors’ contributions

C.-L. Liu and T.-W. Tan designed the study. C.-L. Liu and H.-R. Bi performed the experiments. C.-L. Liu, B. Hu, and L.-H. Fan are involved in the manuscript writing and editing. All the authors read and approved the manuscript.

Funding

This work was supported by the State key laboratory of organicinorganic composites, the National Basic Research Program of China (973 program) (2013CB733600), the National Nature Science Foundation of China (21390202, 21476017).

Compliance with ethical standards

Competing interests

The authors declare that they have no competing interests.

Ethical statement

This article does not contain any studies with human participants performed by any of the authors.

Supplementary material

253_2018_9472_MOESM1_ESM.pdf (443 kb)
ESM 1 (PDF 443 kb)

References

  1. Alonso-Gutierrez J, Kim EM, Batth TS, Cho N, Hu Q, Chan LJ, Petzold CJ, Hillson NJ, Adams PD, Keasling JD, Martin GH, Lee TS (2015) Principal component analysis of proteomics (PCAP) as a tool to direct metabolic engineering. Metab Eng 28:123–133CrossRefGoogle Scholar
  2. Dı’az EE, Stams JMA, Amils R, Sanz LJ (2006) Phenotypic properties and microbial diversity of methanogenic granules from a full-scale upflow anaerobic sludge bed reactor treating brewery wastewater. Appl Environ Microbiol 72(7):4942–4949CrossRefGoogle Scholar
  3. Eisenreich W, Bacher A, Arigoni D, Rohdich F (2004) Biosynthesis of isoprenoids via the non-mevalonate pathway. Cell Mol Life Sci 61:1401–1426CrossRefGoogle Scholar
  4. Henneman L, van Cruchten AG, Denis SW, Amolins MW, Placzek AT, Gibbs RA, Kulik W, Waterham HR (2008) Detection of nonsterol isoprenoids by HPLC-MS/MS. Anal Biochem 383:18–24CrossRefGoogle Scholar
  5. Julsing MK, Rijpkema M, Woerdenbag HJ, Quax WJ, Kayser O (2007) Functional analysis of genes involved in the biosynthesis of isoprene in Bacillus subtilis. Appl Microbiol Biotechnol 75:1377–1384CrossRefGoogle Scholar
  6. Kevin W, George AC, Jain A, Batth TS, Baidoo EEK, Wang G, Adams PD, Petzold CJ, Keasling JD, Lee TS (2014) Correlation analysis of targeted proteins and metabolites to assess and engineer microbial isopentenol production. Biotechnol Bioeng 111:11Google Scholar
  7. Kim J, Wang C, Jang H-J, Cha M-S, Park J-E, Jo S-Y, Choi E-S, Kim S-W (2016) Isoprene production by Escherichia coli through the exogenous mevalonate pathway with reduced formation of fermentation byproducts. Microb Cell Factories 15(1):214CrossRefGoogle Scholar
  8. Kuzma J, Nemecek-Marshall M, Pollock WH, Fall R (1995) Bacteria produce the volatile hydrocarbon isoprene. Curr Microbiol 30:97–103CrossRefGoogle Scholar
  9. Liu C-L, Fan L-H, Liu L, Tan T-W (2014) Combinational biosynthesis of isoprene by engineering the MEP pathway in Escherichia coli. Process Biochem 49:2078–2085CrossRefGoogle Scholar
  10. Liu C-L, Lv Q, Tan T-W (2015) Joint antisense RNA strategies for regulating isoprene production in Escherichia coli. RSC Adv 5:74892–74898CrossRefGoogle Scholar
  11. Pitera DJ, Paddon CJ, Newman JD, Keasling JD (2007) Balancing a heterologous mevalonate pathway for improved isoprenoid production in Escherichia coli. Metab Eng 9:193–207CrossRefGoogle Scholar
  12. Primak YA, Du M, Miller MC, Wells DH, Nielsen AT, Weyler W, Beck ZQ (2011) Characterization of a feedback-resistant mevalonate kinase from the archaeon Methanosarcina mazei. Appl Environ Microbiol 77:7772–7778CrossRefGoogle Scholar
  13. Redding-Johanson AM, Batth TS, Chan R, Krupa R, Szmidt HL, Adams PD, Keasling JD, Lee TS, Mukhopadhyay A, Petzold CJ (2011) Targeted proteomics for metabolic pathway optimization: application to terpene production. Metab Eng 13:194–203CrossRefGoogle Scholar
  14. Rohmer M (1999) A mevalonate-independent route to isopentenyl diphosphate. Compr Nat Products Chem 45–67Google Scholar
  15. Sanchez JJ, Berenguer JA, Calderon V, Herce MD (1991) Spoilage of a bakery product by isoprene-producing molds. Rev Agroquim Technol Aliment 31:4Google Scholar
  16. Sharkey TD, Yeh SS (2001) Isoprene emission from plants. Annu Rev Plant Physiol Plant Mol Biol 52:407–436CrossRefGoogle Scholar
  17. Whited MG, Feher JF, Benko AD, Cervin AM, Chotani KG, McAuliffe CJ, LaDuca JR, Ben-Shoshan AE, Sanford JK (2010) Development of a gas-phase bioprocess for isoprene-monomer production using metabolic pathway engineering. Ind Biotechnol 12:152–163CrossRefGoogle Scholar
  18. Xue J, Ahring BK (2011) Enhancing isoprene production by genetic modification of the 1-deoxy-d-xylulose-5-phosphate pathway in Bacillus subtilis. Appl Environ Microbiol 77:2399–2405CrossRefGoogle Scholar
  19. Yang J, Zhao G, Sun Y, Zheng Y, Jiang X, Liu W, Xian M (2012) Bio-isoprene production using exogenous MVA pathway and isoprene synthase in Escherichia coli. Bioresour Technol 104:642–647CrossRefGoogle Scholar
  20. Yang C, Gao X, Jiang Y, Sun B, Gao F, Yang S (2016) Synergy between methylerythritol phosphate pathway and mevalonate pathway for isoprene production in Escherichia coli. Metab Eng 37:79–91CrossRefGoogle Scholar
  21. Ye L, Lv X, Yu H (2016) Engineering microbes for isoprene production. Metab Eng 38:125–138CrossRefGoogle Scholar
  22. Yoon SH, Lee YM, Kim JE, Lee SH, Lee JH, Kim JY, Jung KH, Shin YC, Keasling JD, Kim SW (2006) Enhanced lycopene production in Escherichia coli engineered to synthesize isopentenyl diphosphate and dimethylallyl diphosphate from mevalonate. Biotechnol Bioeng 94:1025–1032CrossRefGoogle Scholar
  23. Yoon SH, Lee SH, Das A, Ryu HK, Jang HJ, Kim JY, Oh DK, Keasling JD, Kim SW (2009) Combinatorial expression of bacterial whole mevalonate pathway for the production of beta-carotene in E. coli. J Biotechnol 140:218–226CrossRefGoogle Scholar
  24. Zahiri HS, Yoon SH, Keasling JD, Lee SH, Won Kim S, Yoon SC, Shin YC (2006) Coenzyme Q10 production in recombinant Escherichia coli strains engineered with a heterologous decaprenyl diphosphate synthase gene and foreign mevalonate pathway. Metab Eng 8:406–416CrossRefGoogle Scholar
  25. Zurbriggen A, Kirst H, Melis A (2012) Isoprene production via the mevalonic acid pathway in Escherichia coli (Bacteria). BioEnergy Res 5(4):814–828CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.National Energy R&D Center for Biorefinery, College of Life Science and TechnologyBeijing University of Chemical TechnologyBeijingPeople’s Republic of China
  2. 2.National Engineering Laboratory for Cereal Fermentation TechnologyJiangnan UniversityWuxi CityChina

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