期刊论文详细信息
Biotechnology for Biofuels
Engineering E. coli strain for conversion of short chain fatty acids to bioalcohols
Anu Jose Mattam1  Syed Shams Yazdani1 
[1] Synthetic Biology and Biofuels Group, International Centre for Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, 110067 New Delhi, India
关键词: Fermentation;    Butanol;    Butyric acid;    Clostridium acetobutylicum;    Engineered E. coli;   
Others  :  1085030
DOI  :  10.1186/1754-6834-6-128
 received in 2013-05-21, accepted in 2013-09-04,  发布年份 2013
PDF
【 摘 要 】

Background

Recent progress in production of various biofuel precursors and molecules, such as fatty acids, alcohols and alka(e)nes, is a significant step forward for replacing the fossil fuels with renewable fuels. A two-step process, where fatty acids from sugars are produced in the first step and then converted to corresponding biofuel molecules in the second step, seems more viable and attractive at this stage. We have engineered an Escherichia coli strain to take care of the second step for converting short chain fatty acids into corresponding alcohols by using butyrate kinase (Buk), phosphotransbutyrylase (Ptb) and aldehyde/alcohol dehydrogenase (AdhE2) from Clostridium acetobutylicum.

Results

The engineered E. coli was able to convert butyric acid and other short chain fatty acids of chain length C3 to C7 into corresponding alcohols and the efficiency of conversion varied with different E. coli strain type. Glycerol proved to be a better donor of ATP and electron as compared to glucose for converting butyric acid to butanol. The engineered E. coli was able to tolerate up to 100 mM butyric acid and produced butanol with the conversion rate close to 100% under anaerobic condition. Deletion of native genes, such as fumarate reductase (frdA) and alcohol dehydrogenase (adhE), responsible for side products succinate and ethanol, which act as electron sink and could compete with butyric acid uptake, did not improve the butanol production efficiency. Indigenous acyl-CoA synthetase (fadD) was found to play no role in the conversion of butyric acid to butanol. Engineered E. coli was cultivated in a bioreactor under controlled condition where 60 mM butanol was produced within 24 h of cultivation. A continuous bioreactor with the provision of cell recycling allowed the continuous production of butanol at the average productivity of 7.6 mmol/l/h until 240 h.

Conclusions

E. coli engineered with the pathway from C. acetobutylicum could efficiently convert butyric acid to butanol. Other short chain fatty acids with the chain length of C3 to C7 were also converted to the corresponding alcohols. The ability of engineered strain to convert butyric acid to butanol continuously demonstrates commercial significance of the system.

【 授权许可】

   
2013 Mattam and Yazdani; licensee BioMed Central Ltd.

【 预 览 】
附件列表
Files Size Format View
20150113170150170.pdf 1842KB PDF download
Figure 7. 94KB Image download
Figure 6. 94KB Image download
Figure 5. 54KB Image download
Figure 3. 75KB Image download
Figure 3. 114KB Image download
Figure 2. 61KB Image download
Figure 1. 9KB Image download
【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 3.

Figure 5.

Figure 6.

Figure 7.

【 参考文献 】
  • [1]Bhattarai K, Stalick WM, McKay S, Geme G, Bhattarai N: Biofuel: an alternative to fossil fuel for alleviating world energy and economic crises. J Environ Sci Health A Tox Hazard Subst Environ Eng 2011, 46:1424-1442.
  • [2]Mussatto SI, Dragone G, Guimarães PM, Silva JP, Carneiro LM, Roberto IC, Vicente A, Domingues L, Teixeira JA: Technological trends, global market, and challenges of bio-ethanol production. Biotechnol Adv 2010, 28:817-830.
  • [3]Jin C, Yao M, Liu H, Leed CF, Ji J: Progress in the production and application of n-butanol as a biofuel. Renew Sustain Energy Rev 2011, 15:4080-4106.
  • [4]Green EM: Fermentative production of butanol–the industrial perspective. Curr Opin Biotechnol 2011, 22:337-343.
  • [5]Dürre P: Biobutanol: an attractive biofuel. Biotechnol J 2007, 2:1525-1534.
  • [6]Gheshlaghi R, Scharer JM, Moo-Young M, Chou CP: Metabolic pathways of clostridia for producing butanol. Biotechnol Adv 2009, 27:764-781.
  • [7]Zheng YN, Li LZ, Xian M, Ma YJ, Yang JM, Xu X, He DZ: Problems with the microbial production of butanol. J Ind Microbiol Biotechnol 2009, 36:1127-1138.
  • [8]Atsumi S, Cann AF, Connor MR, Shen CR, Smith KM, Brynildsen MP, Chou KJ, Hanai T, Liao JC: Metabolic engineering of Escherichia coli for 1-butanol production. Metab Eng 2008, 10:305-311.
  • [9]Inui M, Suda M, Kimura S, Yasuda K, Suzuki H, Toda H, Yamamoto S, Okino S, Suzuki N, Yukawa H: Expression of Clostridium acetobutylicum butanol synthetic genes in Escherichia coli. Appl Microbiol Biotechnol 2008, 77:1305-1316.
  • [10]Chen SK, Chin WC, Tsuge K, Huang CC, Li SY: Fermentation approach for enhancing 1-butanol production using engineered butanologenic Escherichia coli. Bioresour Technol 2013, 145:204-209.
  • [11]Steen EJ, Chan R, Prasad N, Myers S, Petzold CJ, Redding A, Ouellet M, Keasling JD: Metabolic engineering of Saccharomyces cerevisiae for the production of n-butanol. Microb Cell Fact 2008, 7:36. BioMed Central Full Text
  • [12]Shen CR, Lan EI, Dekishima Y, Baez A, Cho KM, Liao JC: Driving forces enable high-titer anaerobic 1-butanol synthesis in Escherichia coli. Appl Environ Microbiol 2011, 77:2905-2915.
  • [13]Lennen RM, Pfleger BF: Engineering Escherichia coli to synthesize free fatty acids. Trends Biotechnol 2012, 30:659-667.
  • [14]Steen EJ, Kang Y, Bokinsky G, Hu Z, Schirmer A, McClure A, Del Cardayre SB, Keasling JD: Microbial production of fatty-acid-derived fuels and chemicals from plant biomass. Nature 2010, 463:559-562.
  • [15]Zhang F, Ouellet M, Batth TS, Adams PD, Petzold CJ, Mukhopadhyay A, Keasling JD: Enhancing fatty acid production by the expression of the regulatory transcription factor FadR. Metab Eng 2012, 14:653-660.
  • [16]Zhang C, Yang H, Yang F, Ma Y: Current progress on butyric acid production by fermentation. Curr Microbiol 2009, 59:656-663.
  • [17]Wei D, Liu X, Yang ST: Butyric acid production from sugarcane bagasse hydrolysate by Clostridium tyrobutyricum immobilized in a fibrous-bed bioreactor. Bioresour Technol 2013, 129:553-560.
  • [18]Howard TP, Middelhaufe S, Moore K, Edner C, Kolak DM, Taylor GN, Parker DA, Lee R, Smirnoff N, Aves SJ, Love J: Synthesis of customized petroleum-replica fuel molecules by targeted modification of free fatty acid pools in Escherichia coli. Proc Natl Acad Sci U S A 2013, 110:7636-7641.
  • [19]Akhtar MK, Turner NJ, Jones PR: Carboxylic acid reductase is a versatile enzyme for the conversion of fatty acids into fuels and chemical commodities. Proc Natl Acad Sci U S A 2013, 110:87-92.
  • [20]Richter H, Qureshi N, Heger S, Dien B, Cotta MA, Angenent LT: Prolonged conversion of n-butyrate to n-butanol with Clostridium saccharoperbutylacetonicum in a two-stage continuous culture with in-situ product removal. Biotechnol Bioeng 2012, 109:913-921.
  • [21]Baba S, Tashiro Y, Shinto H, Sonomoto K: Development of high-speed and highly efficient butanol production systems from butyric acid with high density of living cells of Clostridium saccharoperbutylacetonicum. J Biotechnol 2012, 157:605-612.
  • [22]Tashiro Y, Shinto H, Hayashi M, Baba S, Kobayashi G, Sonomoto K: Novel high-efficient butanol production from butyrate by non-growing Clostridium saccharoperbutylacetonicum N1-4 (ATCC 13564) with methyl viologen. J Biosci Bioeng 2007, 104:238-240.
  • [23]Jurgens G, Survase S, Berezina O, Sklavounos E, Linnekoski J, Kurkijärvi A, Väkevä M, van Heiningen A, Granström T: Butanol production from lignocellulosics. Biotechnol Lett 2012, 34:1415-1434.
  • [24]Cary JW, Petersen DJ, Papoutsakis ET, Bennett GN: Cloning and expression of Clostridium acetobutylicum phosphotransbutyrylase and butyrate kinase genes in Escherichia coli. J Bacteriol 1988, 170:4613-4618.
  • [25]Clark SW, Bennett GN, Rudolph FB: Isolation and characterization of mutants of Clostridium acetobutylicum ATCC 824 deficient in acetoacetyl-Coenzyme A:Acetate/Butyrate:Coenzyme A-Transferase (EC 2.8.3.9) and in Other Solvent Pathway Enzymes. Appl Environ Microbiol 1989, 55:970-976.
  • [26]Black PN, DiRusso CC, Metzger AK, Heimert TL: Cloning, sequencing, and expression of the fadD gene of Escherichia coli encoding acyl coenzyme a synthetase. J Biol Chem 1992, 267:25513-25520.
  • [27]Fontaine L, Meynial- Salles I, Girbai L, Yang X, Croux C, Soucaille P: Molecular characterization and transcriptional analysis of adhE2, the gene encoding the NADH – dependent aldehyde/alcohol dehydrogenase responsible for butanol production in alcohologenic cultures of Clostridium acetobutylicum ATCC 824. J Bacteriol 2002, 184:821-830.
  • [28]Hartmanis MG: Butyrate kinase from Clostridium acetobutylicum. J Biol Chem 1987, 262:617-621.
  • [29]Berezina OV, Zakharova NV, Brandt A, Yarotsky SV, Schwarz WH, Zverlov VV: Reconstructing the clostridial n-butanol metabolic pathway in Lactobacillus brevis. Appl Microbiol Biotechnol 2010, 87:635-646.
  • [30]Sambrook J, Fritsch EF, Maniatis T: Molecular cloning, a laboratory manual. 2nd edition. Cold Spring Harbour, NY: Cold Spring Harbor Laboratory Press; 1989.
  • [31]Andersch W, Bahl H, Gottschalk G: Level of enzymes involved in acetate, butyrate, acetone and butanol formation by Clostridium acetobutylicum. Eur J Appl Microbiol Biotechnol 1983, 18:327-333.
  • [32]Kagi J, Vallee B: The role of zinc in alcohol dehydrogenase: the effect of metal binding agents on the structure of the yeast alcohol dehydrogenase molecule. J Biol Chem 1960, 235:3188-3192.
  • [33]Rose IA: Acetate kinase of bacteria (acetokinase). Methods Enzymol 1955, 1:591-593.
  文献评价指标  
  下载次数:77次 浏览次数:19次