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Engineering Klebsiella oxytoca for efficient 2, 3-butanediol production through insertional inactivation of acetaldehyde dehydrogenase gene

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Abstract

Ethanol was a major byproduct of 2,3-butanediol (2,3-BD) fermentation by Klebsiella oxytoca ME-UD-3. In order to achieve a high efficiency of 2,3-BD production, K. oxytoca mutants deficient in ethanol formation were successfully constructed by replace the aldA gene coding for aldehyde dehydrogenase with a tetracycline resistance cassette. The results suggested that inactivation of aldA led to a significantly improved 2,3-BD production. The carbon flux to 2,3-BD was enhanced by eliminating the byproducing ethanol and at the same time reducing the accumulation of another byproduct acetoin. At last, by fed-batch culturing of the mutant, the final 2,3-BD titer up to 130 g/l with the productivity of 1.63 g/l.h and the 2,3-BD yield relative to glucose of 0.48 g/g was obtained.

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References

  • Afschar AS, Bellgardt KH, Vaz Rossell CE, Czok A, Schaller K (1991) The production of 2, 3-butanediol by fermentation of high test molasses. Appl Microbiol Biotechnol 34:582–585

    Article  CAS  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein–dye binding. Ana Biochem 72:248–254

    Article  CAS  Google Scholar 

  • Celińska E, Grajek W (2009) Biotechnological production of 2, 3-butanediol – current state and prospects. Biotechnol Adv. doi:10.1016/j.biotechadv.2009.05.002

    Google Scholar 

  • Cheng KK, Liu HJ, Liu DH (2005) Multiple growth inhibition of Klebsiella pneumoniae in 1, 3-propanediol fermentation. Biotechnol Lett 27:19–22

    Article  CAS  Google Scholar 

  • Converti A, Perego P, Del Borghi M (2003) Effect of specific oxygen uptake rate on Enterobacter aerogenes energetics: carbon and reduction degree balances in batch cultivations. Biotechnol Bioeng 82:370–377

    Article  CAS  Google Scholar 

  • Fournet-Fayard S, Joly B, Forestier C (1995) Transformation of wild type Klebsiella pneumoniae with plasmid DNA by electroporation. J Microbiol Methods 24:49–54

    Article  Google Scholar 

  • Garg SK, Jain A (1995) Fermentative production of 2, 3-butanediol: a review. Bioresour Technol 51:103–109

    Article  CAS  Google Scholar 

  • Huang H, Ji XJ, Li S, Du J, Lian M, Gao Z, Hu N (2007) A high 2,3-butanediol producing Klebsiella oxytoca and its application. Chinese Patent (in Chinese). 200710021641.5

  • Jeong KJ, Lee HS, Lee SY, Chang YK (1998) Efficient transformation of Klebsiella oxytoca by electroporation. Biotechnol Bioprocess Eng 3:48–49

    Article  Google Scholar 

  • Ji XJ, Huang H, Li S, Du J, Lian M (2008) Enhanced 2, 3-butanediol production by altering the mixed acid fermentation pathway in Klebsiella oxytoca. Biotechnol Lett 30:731–734

    Article  CAS  Google Scholar 

  • Ji XJ, Huang H, Du J, Zhu JG, Ren LJ, Hu N, Li S (2009) Enhanced 2, 3-butanediol production by Klebsiella oxytoca using a two-stage agitation speed control strategy. Bioresour Technol 100:3410–3414

    Article  CAS  Google Scholar 

  • Johansen L, Bryn K, Stormer FC (1975) Physiological and biochemical role of the butanediol pathway in Aerobacter (Enterobacter) aerogenes. J Bacteriol 123:1124–1130

    CAS  Google Scholar 

  • Joseph S, David WR (2001) Molecular cloning: a laboratory manual (Third edition). Cold Spring Harbor Laboratory Press, Cold Spring Harbor

    Google Scholar 

  • Kosaric N, Magee RJ, Blaszczyk R (1992) Redox potential measurement for monitoring glucose and xylose conversion by K. pneumoniae. Chem Biochem Eng Q 6:145–152

    CAS  Google Scholar 

  • Ma C, Wang A, Qin J, Li L, Ai X, Jiang T, Tang H, Xu P (2009) Enhanced 2, 3-butanediol production by Klebsiella pneumoniae SDM. Appl Microbiol Biotechnol 82:49–57

    Article  CAS  Google Scholar 

  • Ma LC, Lee SL, Lee CZ, Yin SJ, Wang JT (2005) Molecular cloning and kinetic characterization of an aldehyde dehydrogenase gene in Klebsiella pneumoniae. J Formos Med Assoc 104:221–230

    CAS  Google Scholar 

  • Makino S, Uchida I, Terakado N, Sasakawa C, Yoshikawa M (1989) Molecular characterization and protein analysis of the cap region, which is essential for encapsulation in Bacillus anthracis. J Bacteriol 171:722–730

    CAS  Google Scholar 

  • Qureshi N, Cheryan M (1989) Production of 2, 3-butanediol by Klebsiella oxytoca. Appl Microbiol Biotechnol 30:440–443

    Article  CAS  Google Scholar 

  • Ragauskas AJ, Williams CK, Davison BH, Britovsek G, Cairney J, Eckert CA, Frederick WJ Jr, Hallett JP, Leak DJ, Liotta CL, Mielenz JR, Murphy R, Templer R, Tschaplinski T (2006) The path forward for biofuels and biomaterials. Science 311:484–498

    Article  CAS  Google Scholar 

  • Syu MJ (2001) Biological production of 2, 3-butanediol. Appl Microbiol Biotechnol 55:10–18

    Article  CAS  Google Scholar 

  • Tran AV, Chambers RP (1987) The dehydration of fermentative 2, 3-butanediol into methyl ethyl ketone. Biotechnol Bioeng 29:343–351

    Article  CAS  Google Scholar 

  • van Haveren J, Scott EL, Sanders J (2007) Bulk chemicals from biomass. Biofuels, Bioprod Bioref 2:41–57

    Article  Google Scholar 

  • Wu KJ, Saratale GD, Lo YC, Chen WM, Tseng ZJ, Chang MC, Tsai BC, Su A, Chang JS (2008) Simultaneous production of 2, 3-butanediol, ethanol and hydrogen with a Klebsiella sp. strain isolated from sewage sludge. Bioresour Technol 99:7966–7970

    Article  CAS  Google Scholar 

  • Xu YZ, Guo NN, Zheng ZM, Ou XJ, Liu HJ, Liu DH (2009) Metabolism in 1, 3-propanediol fed-batch fermentation by a D-lactate deficient mutant of Klebsiella pneumoniae. Biotechnol Bioeng. doi:10.1002/bit.22455

    Google Scholar 

  • Yang G, Tian JS, Li JL (2007) Fermentation of 1, 3-propanediol by a lactate deficient mutant of Klebsiella oxytoca under microaerobic conditions. Appl Microbiol Biotechnol 73:1017–1024

    Article  CAS  Google Scholar 

  • Zeng AP, Biebl H (2002) Bulk chemicals from biotechnology: the case of 1, 3-propanediol production and the new trends. Adv Biochem Eng Biotechnol 74:239–259

    CAS  Google Scholar 

  • Zeng AP, Ross A, Biebl H, Tag C, Günzel B, Deckwer WD (1994) Multiple product inhibition and growth modeling of Clostridium butyricum and Klebsiella pneumoniae in glycerol fermentation. Biotechnol Bioeng 44:902–911

    Article  CAS  Google Scholar 

  • Zhang Y, Li Y, Du C, Liu M, Cao Z (2006) Inactivation of aldehyde dehydrogenase: A key factor for engineering 1, 3-propanediol production by Klebsiella pneumoniae. Metab Eng 8:578–586

    Article  CAS  Google Scholar 

  • Zhu JB, Long MN, Xu FC, Wu XB, Xu HJ (2007) Enhanced hydrogen production by insertional inactivation of adhE gene in Klebsiella oxytoca HP1. Chinese Sci Bull 52:492–496

    Article  CAS  Google Scholar 

  • Zhu JG, Li S, Ji XJ, Huang H, Hu N (2009) Enhanced 1, 3-propanediol production in recombinant Klebsiella pneumoniae carrying the gene yqhD encoding 1, 3-propanediol oxidoreductase isoenzyme. World J Microbiol Biotechnol 25:1217–1223

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was financially supported by the National Natural Science Foundation of China (Grant No. 20606018), the Key Program of National Natural Science Foundation of China (Grant No. 20936002), the National Basic Research Program of China (Grant No. 2007CB707805), and the National High Technology Research and Development Program of China (Grant No. 2006AA02Z244). X.-J. Ji was supported by the Innovation Fund for Doctoral Dissertation of Nanjing University of Technology (Grant No. BSCX200808).

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Authors and Affiliations

  1. State Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing University of Technology, No. 5 Xinmofan Road, Nanjing, 210009, People’s Republic of China

    Xiao-Jun Ji, He Huang, Jian-Guo Zhu, Lu-Jing Ren, Zhi-Kui Nie, Jun Du & Shuang Li

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  1. Xiao-Jun Ji
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  2. He Huang
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  3. Jian-Guo Zhu
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  4. Lu-Jing Ren
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  5. Zhi-Kui Nie
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  6. Jun Du
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  7. Shuang Li
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Correspondence to He Huang.

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Ji, XJ., Huang, H., Zhu, JG. et al. Engineering Klebsiella oxytoca for efficient 2, 3-butanediol production through insertional inactivation of acetaldehyde dehydrogenase gene. Appl Microbiol Biotechnol 85, 1751–1758 (2010). https://doi.org/10.1007/s00253-009-2222-2

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  • DOI: https://doi.org/10.1007/s00253-009-2222-2

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