Skip to main content
SpringerLink
Log in

Metabolic engineering of Escherichia coli for production of biodiesel from fatty alcohols and acetyl-CoA

  • Bioenergy and biofuels
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

Microbial production of biodiesel from renewable feedstock has attracted intensive attention. Biodiesel is known to be produced from short-chain alcohols and fatty acyl-CoAs through the expression of wax ester synthase/fatty acyl-CoA: diacylglycerol acyltransferase that catalyzes the esterification of short-chain alcohols and fatty acyl-CoAs. Here, we engineered Escherichia coli to produce various fatty alcohol acetate esters, which depend on the expression of Saccharomyces cerevisiae alcohol acetyltransferase ATF1 that catalyzes the esterification of fatty alcohols and acetyl-CoA. The fatty acid biosynthetic pathways generate fatty acyl-ACPs, fatty acyl-CoAs, or fatty acids, which can be converted to fatty alcohols by fatty acyl-CoA reductase, fatty acyl-ACP reductase, or carboxylic acid reductase, respectively. This study showed the biosynthesis of biodiesel from three fatty acid biosynthetic pathway intermediates.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2

Similar content being viewed by others

References

  • Akhtar MK, Turner NJ, Jones PR (2013) Carboxylic acid reductase is a versatile enzyme for the conversion of fatty acids into fuels and chemical commodities. Proc Natl Acad Sci 110(1):87–92

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Atsumi S, Hanai T, Liao JC (2008) Non-fermentative pathways for synthesis of branched-chain higher alcohols as biofuels. Nature 451(7174):86–89

    Article  CAS  PubMed  Google Scholar 

  • Avalos JL, Fink GR, Stephanopoulos G (2013) Compartmentalization of metabolic pathways in yeast mitochondria improves the production of branched-chain alcohols. Nat Biotechnol 31:335–341

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Barnes EM, Swindell A, Wakil SJ (1970) Purification and properties of a palmityl thioesterase II from Escherichia coli. J Biol Chem 245(12):3122–3128

    CAS  PubMed  Google Scholar 

  • Barney BM (2014) Metabolic engineering: the sweet smell of biosynthesis. Nat Chem Biol 10(4):246–247

    Article  CAS  PubMed  Google Scholar 

  • Bokinsky G, Peralta-Yahya PP, George A, Holmes BM, Steen EJ, Dietrich J, Lee TS, Tullman-Ercek D, Voigt CA, Simmons BA (2011) Synthesis of three advanced biofuels from ionic liquid-pretreated switchgrass using engineered Escherichia coli. Proc Natl Acad Sci 108(50):19949–19954

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Caspeta L, Nielsen J (2013) Economic and environmental impacts of microbial biodiesel. Nat Biotechnol 31(9):789–793

    Article  CAS  PubMed  Google Scholar 

  • Choi YJ, Lee SY (2013) Microbial production of short-chain alkanes. Nature 502(7472):571–574

    Article  CAS  PubMed  Google Scholar 

  • Datsenko KA, Wanner BL (2000) One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci 97(12):6640–6645

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Davis MS, Solbiati J, Cronan JE (2000) Overproduction of acetyl-CoA carboxylase activity increases the rate of fatty acid biosynthesis in Escherichia coli. J Biol Chem 275(37):28593–28598

    Article  CAS  PubMed  Google Scholar 

  • Duan Y, Zhu Z, Cai K, Tan X, Lu X (2011) De novo biosynthesis of biodiesel by Escherichia coli in optimized fed-batch cultivation. PLoS One 6(5):e20265

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Elbahloul Y, Steinbüchel A (2010) Pilot-scale production of fatty acid ethyl esters by an engineered Escherichia coli strain harboring the p (Microdiesel) plasmid. Appl Environ Microbiol 76(13):4560–4565

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Fujii T, Nagasawa N, Iwamatsu A, Bogaki T, Tamai Y, Hamachi M (1994) Molecular cloning, sequence analysis, and expression of the yeast alcohol acetyltransferase gene. Appl Environ Microbiol 60(8):2786–2792

    PubMed Central  CAS  PubMed  Google Scholar 

  • Fukuda H, Kondo A, Noda H (2001) Biodiesel fuel production by transesterification of oils. J Biosci Bioeng 92(5):405–416

    Article  CAS  PubMed  Google Scholar 

  • Gu S-H, Wu K-M, Guo Y-Y, Pickett JA, Field LM, Zhou J-J, Zhang Y-J (2013) Identification of genes expressed in the sex pheromone gland of the black cutworm Agrotis ipsilon with putative roles in sex pheromone biosynthesis and transport. BMC Genomics 14(1):636

  • Guo D, Zhu J, Deng Z, Liu T (2014) Metabolic engineering of Escherichia coli for production of fatty acid short-chain esters through combination of the fatty acid and 2-keto acid pathways. Metab Eng 22:69–75

    Article  CAS  PubMed  Google Scholar 

  • Hanai T, Atsumi S, Liao J (2007) Engineered synthetic pathway for isopropanol production in Escherichia coli. Appl Environ Microbiol 73(24):7814–7818

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Handke P, Lynch SA, Gill RT (2011) Application and engineering of fatty acid biosynthesis in Escherichia coli for advanced fuels and chemicals. Metab Eng 13(1):28–37

    Article  CAS  PubMed  Google Scholar 

  • Hofvander P, Doan TT, Hamberg M (2011) A prokaryotic acyl-CoA reductase performing reduction of fatty acyl-CoA to fatty alcohol. FEBS Lett 585(22):3538–3543

    Article  CAS  PubMed  Google Scholar 

  • Howard TP, Middelhaufe S, Moore K, Edner C, Kolak DM, Taylor GN, Parker DA, Lee R, Smirnoff N, Aves SJ (2013) Synthesis of customized petroleum-replica fuel molecules by targeted modification of free fatty acid pools in Escherichia coli. Proc Natl Acad Sci 110(19):7636–7641

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Janßen HJ, Steinbuchel A (2014) Fatty acid synthesis in Escherichia coli and its applications towards the production of fatty acid based biofuels. Biotechnol Biofuels 7(7)

  • Kalscheuer R, Steinbüchel A (2003) A novel bifunctional wax ester synthase/acyl-CoA: diacylglycerol acyltransferase mediates wax ester and triacylglycerol biosynthesis inAcinetobacter calcoaceticus ADP1. J Biol Chem 278(10):8075–8082

    Article  CAS  PubMed  Google Scholar 

  • Kalscheuer R, Luftmann H, Steinbüchel A (2004) Synthesis of novel lipids in Saccharomyces cerevisiae by heterologous expression of an unspecific bacterial acyltransferase. Appl Environ Microbiol 70(12):7119–7125

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Kalscheuer R, Stölting T, Steinbüchel A (2006) Microdiesel: Escherichia coli engineered for fuel production. Microbiology 152(9):2529–2536

    Article  CAS  PubMed  Google Scholar 

  • Layton DS, Trinh CT (2014) Engineering modular ester fermentative pathways in Escherichia coli. Metab Eng 26:77–88

    Article  CAS  Google Scholar 

  • Lenneman EM, Ohlert JM, Palani NP, Bm B (2013) Fatty alcohols for wax esters in Marinobacter aquaeolei VT8: two optional routes in the wax biosynthesis pathway. Appl Environ Microbiol 79(22):7055–7062

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Leung DYC, Wu X, Leung M (2010) A review on biodiesel production using catalyzed transesterification. Appl Energy 87(4):1083–1095

    Article  CAS  Google Scholar 

  • Lilly M, Lambrechts M, Pretorius I (2000) Effect of increased yeast alcohol acetyltransferase activity on flavor profiles of wine and distillates. Appl Environ Microbiol 66(2):744–753

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Liu A, Tan X, Yao L, Lu X (2013) Fatty alcohol production in engineered E. coli expressing marinobacter fatty acyl-CoA reductases. Appl Microbiol Biotechnol 97(15):7061–7071

    Article  CAS  PubMed  Google Scholar 

  • Liu R, Zhu F, Lu L, Fu A, Lu J, Deng Z, Liu T (2014) Metabolic engineering of fatty acyl-ACP reductase-dependent pathway to improve fatty alcohol production in Escherichia coli. Metab Eng 22:10–21

    Article  CAS  PubMed  Google Scholar 

  • Lu X, Vora H, Khosla C (2008) Overproduction of free fatty acids in E. coli: implications for biodiesel production. Metab Eng 10(6):333–339

    Article  CAS  PubMed  Google Scholar 

  • Ma F, Hanna MA (1999) Biodiesel production: a review. Bioresour Technol 70(1):1–15

    Article  CAS  Google Scholar 

  • Magnuson K, Jackowski S, Rock C, Cronan J (1993) Regulation of fatty acid biosynthesis in Escherichia coli. Microbiol Rev 57(3):522–542

    PubMed Central  CAS  PubMed  Google Scholar 

  • Mason AB, Dufour JP (2000) Alcohol acetyltransferases and the significance of ester synthesis in yeast. Yeast 16(14):1287–1298

    Article  CAS  PubMed  Google Scholar 

  • Mata TM, Martins AA, Caetano NS (2010) Microalgae for biodiesel production and other applications: a review. Renew Sust Energ Rev 14(1):217–232

    Article  CAS  Google Scholar 

  • Menendez-Bravo S, Comba S, Sabatini M, Arabolaza A, Gramajo H (2014) Expanding the chemical diversity of natural esters by engineering a polyketide-derived pathway into Escherichia coli. Metab Eng 24:97–106

    Article  CAS  PubMed  Google Scholar 

  • Reiser S, Somerville C (1997) Isolation of mutants of Acinetobacter calcoaceticus deficient in wax ester synthesis and complementation of one mutation with a gene encoding a fatty acyl coenzyme a reductase. J Bacteriol 179(9):2969–2975

    PubMed Central  CAS  PubMed  Google Scholar 

  • Rodriguez GM, Tashiro Y, Atsumi S (2014) Expanding ester biosynthesis in Escherichia coli. Nat Chem Biol 10(4):259–265

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Runguphan W, Keasling JD (2014) Metabolic engineering of Saccharomyces cerevisiae for production of fatty acid-derived biofuels and chemicals. Metab Eng 21:103–113

    Article  CAS  PubMed  Google Scholar 

  • Saerens SM, Delvaux FR, Verstrepen KJ, Thevelein JM (2010) Production and biological function of volatile esters in Saccharomyces cerevisiae. Microb Biotechnol 3(2):165–177

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Schirmer A, Rude MA, Li X, Popova E, Del Cardayre SB (2010) Microbial biosynthesis of alkanes. Science 329(5991):559–562

    Article  CAS  PubMed  Google Scholar 

  • Shi S, Valle-Rodríguez JO, Khoomrung S, Siewers V, Nielsen J (2012) Functional expression and characterization of five wax ester synthases in Saccharomyces cerevisiae and their utility for biodiesel production. Biotechnol Biofuels 5(3):7

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Steen EJ, Kang Y, Bokinsky G, Hu Z, Schirmer A, McClure A, Del Cardayre SB, Keasling JD (2010) Microbial production of fatty-acid-derived fuels and chemicals from plant biomass. Nature 463(7280):559–562

    Article  CAS  PubMed  Google Scholar 

  • Stephanopoulos G (2007) Challenges in engineering microbes for biofuels production. Science 315(5813):801–804

    Article  CAS  PubMed  Google Scholar 

  • Tai Y-S, Xiong M, Zhang K (2015) Engineered biosynthesis of medium-chain esters in Escherichia coli. Metab Eng 27:20–28

    Article  CAS  PubMed  Google Scholar 

  • Thompson B, Moon TS, Nielsen DR (2014) ‘Hybrid’processing strategies for expanding and improving the synthesis of renewable bioproducts. Curr Opin Biotechnol 30:17–23

    Article  CAS  PubMed  Google Scholar 

  • Valle-Rodríguez JO, Shi S, Siewers V, Nielsen J (2014) Metabolic engineering of Saccharomyces cerevisiae for production of fatty acid ethyl esters, an advanced biofuel, by eliminating non-essential fatty acid utilization pathways. Appl Energy 115:226–232

    Article  Google Scholar 

  • Varman AM, Xiao Y, Pakrasi HB, Tang YJ (2013) Metabolic engineering of Synechocystis sp. strain PCC 6803 for isobutanol production. Appl Environ Microbiol 79(3):908–914

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Verstrepen KJ, Van Laere SD, Vanderhaegen BM, Derdelinckx G, Dufour J-P, Pretorius IS, Winderickx J, Thevelein JM, Delvaux FR (2003) Expression levels of the yeast alcohol acetyltransferase genes ATF1, Lg-ATF1, and ATF2 control the formation of a broad range of volatile esters. Appl Environ Microbiol 69(9):5228–5237

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Wahlen BD, Oswald WS, Seefeldt LC, Barney BM (2009) Purification, characterization, and potential bacterial wax production role of an NADPH-dependent fatty aldehyde reductase from Marinobacter aquaeolei VT8. Appl Environ Microbiol 75(9):2758–2764

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Willis RM, Wahlen BD, Seefeldt LC, Barney BM (2011) Characterization of a fatty acyl-CoA reductase from Marinobacter aquaeolei VT8: a bacterial enzyme catalyzing the reduction of fatty acyl-CoA to fatty alcohol. Biochemistry 50(48):10550–10558

    Article  CAS  PubMed  Google Scholar 

  • Yoshimoto H, Fujiwara D, Momma T, Ito C, Sone H, Kaneko Y, Tamai Y (1998) Characterization of the ATF1 and Lg-ATF1 genes encoding alcohol acetyltransferases in the bottom fermenting yeast Saccharomyces pastorianus. J Ferment Bioeng 86(1):15–20

    Article  CAS  Google Scholar 

  • Yoshimoto H, Fukushige T, Yonezawa T, Sone H (2002) Genetic and physiological analysis of branched-chain alcohols and isoamyl acetate production in Saccharomyces cerevisiae. Appl Microbiol Biotechnol 59(4–5):501–508

    CAS  PubMed  Google Scholar 

  • Yu X, Liu T, Zhu F, Khosla C (2011) In vitro reconstitution and steady-state analysis of the fatty acid synthase from Escherichia coli. Proc Natl Acad Sci 108(46):18643–18648

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Zhang F, Carothers JM, Keasling JD (2012) Design of a dynamic sensor-regulator system for production of chemicals and fuels derived from fatty acids. Nat Biotechnol 30(4):354–359

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by National Nature Science Foundation of China (50968002, 50727803, and 30860073), Nature Science Foundation of Jiangxi province (2014BAB203011, 2008GQY0102), and Foundation of Jiangxi Educational Committee (GJJ10236).

Conflict of interest

The authors declare that they have no competing interests.

Author information

Authors and Affiliations

  1. College of Life and Environmental Sciences, Gannan Normal University, Ganzhou, People’s Republic of China

    Daoyi Guo

  2. Key Laboratory of Organo-Pharmaceutical Chemistry, Gannan Normal University, Jiangxi Province, Ganzhou, 341000, People’s Republic of China

    Daoyi Guo, Hong Pan & Xun Li

Authors
  1. Daoyi Guo
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hong Pan
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Xun Li
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Daoyi Guo or Xun Li.

Electronic supplementary material

ESM 1

(PDF 460 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Guo, D., Pan, H. & Li, X. Metabolic engineering of Escherichia coli for production of biodiesel from fatty alcohols and acetyl-CoA. Appl Microbiol Biotechnol 99, 7805–7812 (2015). https://doi.org/10.1007/s00253-015-6809-5

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-015-6809-5

Keywords

Search

Navigation