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Improved cell growth and biosynthesis of glycolic acid by overexpression of membrane-bound pyridine nucleotide transhydrogenase

  • Metabolic Engineering and Synthetic Biology - Original Paper
  • Published:
Journal of Industrial Microbiology & Biotechnology

Abstract

The non-conventional d-xylose metabolism called the Dahms pathway which only requires the expression of at least three enzymes to produce pyruvate and glycolaldehyde has been previously engineered in Escherichia coli. Strains that rely on this pathway exhibit lower growth rates which were initially attributed to the perturbed redox homeostasis as evidenced by the lower intracellular NADPH concentrations during exponential growth phase. NADPH-regenerating systems were then tested to restore the redox homeostasis. The membrane-bound pyridine nucleotide transhydrogenase, PntAB, was overexpressed and resulted to a significant increase in biomass and glycolic acid titer and yield. Furthermore, expression of PntAB in an optimized glycolic acid-producing strain improved the growth and product titer significantly. This work demonstrated that compensating for the NADPH demand can be achieved by overexpression of PntAB in E. coli strains assimilating d-xylose through the Dahms pathway. Consequently, increase in biomass accumulation and product concentration was also observed.

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References

  1. Andersen KB, von Meyenburg K (1977) Charges of nicotinamide adenine nucleotides and adenylate energy charge as regulatory parameters of the metabolism in Escherichia coli. J Biol Chem 252:4151–4156

    CAS  PubMed  Google Scholar 

  2. Balzer GJ, Thakker C, Bennett GN, San K-Y (2013) Metabolic engineering of Escherichia coli to minimize byproduct formate and improving succinate productivity through increasing NADH availability by heterologous expression of NAD+-dependent formate dehydrogenase. Metab Eng 20:1–8. https://doi.org/10.1016/j.ymben.2013.07.005

    Article  CAS  PubMed  Google Scholar 

  3. Bologna FP, Andreo CS, Drincovich MF (2007) Escherichia coli malic enzymes: two isoforms with substantial differences in kinetic properties, metabolic regulation, and structure. J Bacteriol 189:5937–5946. https://doi.org/10.1128/JB.00428-07

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Cabulong RB, Lee W-K, Bañares AB et al (2018) Engineering Escherichia coli for glycolic acid production from d-xylose through the Dahms pathway and glyoxylate bypass. Appl Microbiol Biotechnol 102:2179–2189. https://doi.org/10.1007/s00253-018-8744-8

    Article  CAS  PubMed  Google Scholar 

  5. Cabulong RB, Valdehuesa KNG, Ramos KRM et al (2017) Enhanced yield of ethylene glycol production from d-xylose by pathway optimization in Escherichia coli. Enzym Microb Technol 97:11–20. https://doi.org/10.1016/j.enzmictec.2016.10.020

    Article  CAS  Google Scholar 

  6. Choi SY, Park SJ, Kim WJ et al (2016) One-step fermentative production of poly(lactate-co-glycolate) from carbohydrates in Escherichia coli. Nat Biotechnol 34:435–440. https://doi.org/10.1038/nbt.3485

    Article  CAS  PubMed  Google Scholar 

  7. Cui Y-Y, Ling C, Zhang Y-Y et al (2014) Production of shikimic acid from Escherichia coli through chemically inducible chromosomal evolution and cofactor metabolic engineering. Microb Cell Fact 13:21. https://doi.org/10.1186/1475-2859-13-21

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Dahms AS (1974) 3-Deoxy-d-pentulosonic acid aldolase and its role in a new pathway of d-xylose degradation. Biochem Biophys Res Comm 60:1433–1439. https://doi.org/10.1016/0006-291X(74)90358-1

    Article  CAS  PubMed  Google Scholar 

  9. Green MR, Sambrook J (2012) Molecular cloning: a laboratory manual, 4th edn. Cold Spring Harbor Laboratory Press, New York

    Google Scholar 

  10. Haverkorn van Rijsewijk BRB, Kochanowski K, Heinemann M, Sauer U (2016) Distinct transcriptional regulation of the two Escherichia coli transhydrogenases PntAB and UdhA. Microbiol 162:1672–1679. https://doi.org/10.1099/mic.0.000346

    Article  CAS  Google Scholar 

  11. Holm AK, Blank LM, Oldiges M et al (2010) Metabolic and transcriptional response to cofactor perturbations in Escherichia coli. J Biol Chem 285:17498–17506. https://doi.org/10.1074/jbc.M109.095570

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Khattab SMR, Saimura M, Kodaki T (2013) Boost in bioethanol production using recombinant Saccharomyces cerevisiae with mutated strictly NADPH-dependent xylose reductase and NADP+-dependent xylitol dehydrogenase. J Biotechnol 165:153–156. https://doi.org/10.1016/j.jbiotec.2013.03.009

    Article  CAS  PubMed  Google Scholar 

  13. Lee W-H, Kim M-D, Jin Y-S, Seo J-H (2013) Engineering of NADPH regenerators in Escherichia coli for enhanced biotransformation. Appl Microbiol Biotechnol 97:2761–2772. https://doi.org/10.1007/s00253-013-4750-z

    Article  CAS  PubMed  Google Scholar 

  14. Lim JH, Seo SW, Kim SY, Jung GY (2013) Model-driven rebalancing of the intracellular redox state for optimization of a heterologous n-butanol pathway in Escherichia coli. Metab Eng 20:56–62. https://doi.org/10.1016/j.ymben.2013.09.003

    Article  CAS  PubMed  Google Scholar 

  15. Lipson DA (2015) The complex relationship between microbial growth rate and yield and its implications for ecosystem processes. Front Microbiol 6:615. https://doi.org/10.3389/fmicb.2015.00615

    Article  PubMed  PubMed Central  Google Scholar 

  16. Liu H, Ramos KRM, Valdehuesa KNG et al (2013) Biosynthesis of ethylene glycol in Escherichia coli. Appl Microbiol Biotechnol 97:3409–3417. https://doi.org/10.1007/s00253-012-4618-7

    Article  CAS  PubMed  Google Scholar 

  17. Liu J, Li H, Zhao G et al (2018) Redox cofactor engineering in industrial microorganisms: strategies, recent applications and future directions. J Ind Microbiol Biotechnol 45:313–327. https://doi.org/10.1007/s10295-018-2031-7

    Article  CAS  PubMed  Google Scholar 

  18. Meijnen J-P, de Winde JH, Ruijssenaars HJ (2009) Establishment of oxidative d-xylose metabolism in Pseudomonas putida S12. Appl Environ Microbiol 75:2784–2791. https://doi.org/10.1128/AEM.02713-08

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Radek A, Krumbach K, Gätgens J et al (2014) Engineering of Corynebacterium glutamicum for minimized carbon loss during utilization of d-xylose containing substrates. J Biotechnol 192(Pt A):156–160. https://doi.org/10.1016/j.jbiotec.2014.09.026

    Article  CAS  PubMed  Google Scholar 

  20. Rathnasingh C, Raj SM, Lee Y et al (2012) Production of 3-hydroxypropionic acid via malonyl-CoA pathway using recombinant Escherichia coli strains. J Biotechnol 157:633–640. https://doi.org/10.1016/j.jbiotec.2011.06.008

    Article  CAS  PubMed  Google Scholar 

  21. Salusjärvi L, Toivari M, Vehkomäki M-L et al (2017) Production of ethylene glycol or glycolic acid from d-xylose in Saccharomyces cerevisiae. Appl Microbiol Biotechnol 101:8151–8163. https://doi.org/10.1007/s00253-017-8547-3

    Article  CAS  PubMed  Google Scholar 

  22. Sauer U, Canonaco F, Heri S et al (2004) The soluble and membrane-bound transhydrogenases UdhA and PntAB have divergent functions in NADPH metabolism of Escherichia coli. J Biol Chem 279:6613–6619. https://doi.org/10.1074/jbc.M311657200

    Article  CAS  PubMed  Google Scholar 

  23. Shi A, Zhu X, Lu J et al (2013) Activating transhydrogenase and NAD kinase in combination for improving isobutanol production. Metab Eng 16:1–10. https://doi.org/10.1016/j.ymben.2012.11.008

    Article  CAS  PubMed  Google Scholar 

  24. Spaans SK, Weusthuis RA, van der Oost J, Kengen SWM (2015) NADPH-generating systems in bacteria and archaea. Front Microbiol 6:159. https://doi.org/10.3389/fmicb.2015.00742

    Article  Google Scholar 

  25. Stockland AE, San Clemente CL (1969) Multiple forms of lactate dehydrogenase in Staphylococcus aureus. J Bacteriol 100:347–353

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Valdehuesa KNG, Lee W-K, Ramos KRM et al (2015) Identification of aldehyde reductase catalyzing the terminal step for conversion of xylose to butanetriol in engineered Escherichia coli. Bioproc Biosyst Eng 38:1761–1772. https://doi.org/10.1007/s00449-015-1417-4

    Article  CAS  Google Scholar 

  27. Valdehuesa KNG, Liu H, Ramos KRM et al (2014) Direct bioconversion of d-xylose to 1,2,4-butanetriol in an engineered Escherichia coli. Proc Biochem 49:25–32. https://doi.org/10.1016/j.procbio.2013.10.002

    Article  CAS  Google Scholar 

  28. Valdehuesa KNG, Ramos KRM, Nisola GM et al (2018) Everyone loves an underdog: metabolic engineering of the xylose oxidative pathway in recombinant microorganisms. Appl Microbiol Biotechnol 102:7703–7716. https://doi.org/10.1007/s00253-018-9186-z

    Article  CAS  PubMed  Google Scholar 

  29. Wang Y, San K-Y, Bennett GN (2013) Cofactor engineering for advancing chemical biotechnology. Curr Opin Biotechnol 24:994–999. https://doi.org/10.1016/j.copbio.2013.03.022

    Article  CAS  PubMed  Google Scholar 

  30. Wang Y, San K-Y, Bennett GN (2013) Improvement of NADPH bioavailability in Escherichia coli by replacing NAD+-dependent glyceraldehyde-3-phosphate dehydrogenase GapA with NADP+-dependent GapB from Bacillus subtilis and addition of NAD kinase. J Ind Microbiol Biotechnol 40:1449–1460. https://doi.org/10.1007/s10295-013-1335-x

    Article  CAS  PubMed  Google Scholar 

  31. Wasserstrom L, Portugal-Nunes D, Almqvist H et al (2018) Exploring d-xylose oxidation in Saccharomyces cerevisiae through the Weimberg pathway. AMB Express 8:33. https://doi.org/10.1186/s13568-018-0564-9

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Weimberg R (1961) Pentose oxidation by Pseudomonas fragi. J Biol Chem 236:629–635

    CAS  PubMed  Google Scholar 

  33. Xu P, Vansiri A, Bhan N, Koffas MAG (2012) ePathBrick: a synthetic biology platform for engineering metabolic pathways in E. coli. ACS Synth Biol 1:256–266. https://doi.org/10.1021/sb300016b

    Article  CAS  PubMed  Google Scholar 

  34. Zhao C, Zhao Q, Li Y, Zhang Y (2017) Engineering redox homeostasis to develop efficient alcohol-producing microbial cell factories. Microb Cell Fact 16:115. https://doi.org/10.1186/s12934-017-0728-3

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Zhao H, van der Donk WA (2003) Regeneration of cofactors for use in biocatalysis. Curr Opin Biotechnol 14:583–589. https://doi.org/10.1016/j.copbio.2003.09.007

    Article  CAS  PubMed  Google Scholar 

Download references

Funding

This work was supported by Korea Research Fellowship Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT (2015H1D3A1062172 and 2016R1C1B1013252) and by the Ministry of Education (No. 2018R1D1A1B07043993 and No. 2009-0093816).

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Correspondence to Won-Keun Lee or Wook-Jin Chung.

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Cabulong, R.B., Valdehuesa, K.N.G., Bañares, A.B. et al. Improved cell growth and biosynthesis of glycolic acid by overexpression of membrane-bound pyridine nucleotide transhydrogenase. J Ind Microbiol Biotechnol 46, 159–169 (2019). https://doi.org/10.1007/s10295-018-2117-2

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  • DOI: https://doi.org/10.1007/s10295-018-2117-2

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