Biotechnology for Biofuels (Jan 2019)

Methanol fermentation increases the production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli

  • Xin Wang,
  • Xuelin Wang,
  • Xiaolu Lu,
  • Chen Ma,
  • Kequan Chen,
  • Pingkai Ouyang

DOI
https://doi.org/10.1186/s13068-019-1356-4
Journal volume & issue
Vol. 12, no. 1
pp. 1 – 11

Abstract

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Abstract Background Methanol has attracted increased attention as a non-food alternative carbon source to sugar for biological production of chemicals and fuels. Moreover, the high degree of reduction of methanol offers some advantages in increasing the production yields of NAD(P)H-dependent metabolites. Here, we demonstrate an example of methanol bioconversion with the aim of improving production of NAD(P)H-dependent chemicals in synthetic methylotrophic Escherichia coli. Results A synthetic methylotrophic E. coli was engineered with a nicotinamide adenine dinucleotide (NAD+)-dependent methanol dehydrogenase (MDH) and ribulose monophosphate (RuMP) pathway. Regarding the limited MDH activity, the role of activator proteins in vivo was investigated, and the NudF protein was identified capable of improving MDH activity and triggering increased methanol metabolism. Using 13C-methanol-labeling experiments, we confirmed methanol assimilation in the methylotrophic E. coli. A cycling RuMP pathway for methanol assimilation was also demonstrated by detecting multiple labeled carbons for several compounds. Finally, using the NAD(P)H-dependent metabolite lysine as a test, the potential of methanol bioconversion to generate value-added metabolites was determined. To further characterize the benefit of methanol as the carbon source, extra NADH from methanol oxidation was engineered to generate NADPH to improve lysine biosynthesis by expression of the POS5 gene from Saccharomyces cerevisiae, which resulted in a twofold improvement of lysine production. Moreover, this new sink further pulled upstream methanol utilization. Conclusion Through engineering methanol metabolism, lysine biosynthesis, and NADPH regeneration pathway from NADH, the bioconversion of methanol to improve chemical synthesis was successfully achieved in methylotrophic E. coli.

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