Co-production of hydrogen and ethanol by pfkA-deficient Escherichia coli with activated pentose-phosphate pathway: reduction of pyruvate accumulation.

Balaji Sundara Sekar,Subramanian Mohan Raj,Eunhee Seol,Sunghoon Park

doi:10.1186/s13068-016-0510-5

Balaji Sundara Sekar, Subramanian Mohan Raj + Show 2 more

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https://doi.org/10.1186/s13068-016-0510-5

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Abstract

BackgroundFermentative hydrogen (H2) production suffers from low carbon-to-H2 yield, to which problem, co-production of ethanol and H2 has been proposed as a solution. For improved co-production of H2 and ethanol, we developed Escherichia coli BW25113 ΔhycA ΔhyaAB ΔhybBC ΔldhA ΔfrdAB Δpta-ackA ΔpfkA (SH8*) and overexpressed Zwf and Gnd, the key enzymes in the pentose-phosphate (PP) pathway (SH8*_ZG). However, the amount of accumulated pyruvate, which was significant (typically 0.20 mol mol−1 glucose), reduced the co-production yield.ResultsIn this study, as a means of reducing pyruvate accumulation and improving co-production of H2 and ethanol, we developed and studied E. coli SH9*_ZG with functional acetate production pathway for conversion of acetyl-CoA to acetate (pta-ackA+). Our results indicated that the presence of the acetate pathway completely eliminated pyruvate accumulation and substantially improved the co-production of H2 and ethanol, enabling yields of 1.88 and 1.40 mol, respectively, from 1 mol glucose. These yields, significantly, are close to the theoretical maximums of 1.67 mol H2 and 1.67 mol ethanol. To better understand the glycolytic flux distribution, glycolytic flux prediction and RT-PCR analyses were performed.ConclusionThe presence of the acetate pathway along with activation of the PP pathway eliminated pyruvate accumulation, thereby significantly improving co-production of H2 and ethanol. Our strategy is applicable to anaerobic production of biofuels and biochemicals, both of which processes demand high NAD(P)H.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0510-5) contains supplementary material, which is available to authorized users.

Highlights

Fermentative hydrogen (H2) production suffers from low carbon-to-H2 yield, to which problem, coproduction of ethanol and H2 has been proposed as a solution
As a means of eliminating pyruvate accumulation and improving co-production yields, we developed a new E. coli mutant with an intact acetate production pathway (E. coli BW25113 ΔhycA ΔhyaAB ΔhybBC ΔldhA ΔfrdAB ΔpfkA) from SH5 and evolved the strain (SH9*) for growth under anaerobic conditions
It is believed that this promoterregion mutation increased the transcription of pfkB, allowing SH9* to metabolize glucose through the EMP pathway and support cell growth (Fig. 2c)

Summary

Introduction

Fermentative hydrogen (H2) production suffers from low carbon-to-H2 yield, to which problem, coproduction of ethanol and H2 has been proposed as a solution. The amount of accumulated pyruvate, which was significant (typically 0.20 mol mol−1 glucose), reduced the co-production yield. Biological H2 production can be accomplished via dark fermentation, photo-fermentation, or biophotolysis. As an alternative solution to the introduction of heterologous pathways or hybrid process development, we have suggested co-production of H2 and ethanol in a simple, single-reactor system [13]. From glycerol which is a more reduced substrate than glucose, co-production of H2 and ethanol by E. coli [14] and Klebsiella sp. Co-production of H2 and acetaldehyde with glucose as carbon source has been reported. In this case, acetaldehyde should be chemically reduced to ethanol to be used as fuel [16]

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