Metabolic Engineering of Fusarium oxysporum to Improve Its Ethanol-Producing Capability.

George E. Anasontzis,Paul Christakopoulos,Elisavet Kourtoglou,Silas G. Villas-Boâs,Dimitris G. Hatzinikolaou

doi:10.3389/fmicb.2016.00632

Abstract

Fusarium oxysporum is one of the few filamentous fungi capable of fermenting ethanol directly from plant cell wall biomass. It has the enzymatic toolbox necessary to break down biomass to its monosaccharides and, under anaerobic and microaerobic conditions, ferments them to ethanol. Although these traits could enable its use in consolidated processes and thus bypass some of the bottlenecks encountered in ethanol production from lignocellulosic material when Saccharomyces cerevisiae is used—namely its inability to degrade lignocellulose and to consume pentoses—two major disadvantages of F. oxysporum compared to the yeast—its low growth rate and low ethanol productivity—hinder the further development of this process. We had previously identified phosphoglucomutase and transaldolase, two major enzymes of glucose catabolism and the pentose phosphate pathway, as possible bottlenecks in the metabolism of the fungus and we had reported the effect of their constitutive production on the growth characteristics of the fungus. In this study, we investigated the effect of their constitutive production on ethanol productivity under anaerobic conditions. We report an increase in ethanol yield and a concomitant decrease in acetic acid production. Metabolomics analysis revealed that the genetic modifications applied did not simply accelerate the metabolic rate of the microorganism; they also affected the relative concentrations of the various metabolites suggesting an increased channeling toward the chorismate pathway, an activation of the γ-aminobutyric acid shunt, and an excess in NADPH regeneration.

Highlights

Lignocellulose is the complex matrix of cellulose, hemicellulose, and lignin that constitutes the plant cell wall
Considering the perspective of using F. oxysporum for ethanol production from lignocellulose in a consolidated bioprocess, we evaluated the effect of constitutive expression of the respective genes on ethanol productivity using glucose as the sole carbon source
FF11 produced more than 20 g.L−1 of ethanol after 139 h of anaerobic fermentation, while the wild type seemed to decrease its glucose consumption rate at around 120 h, with an ethanol concentration of less than 10 g L−1

Summary

Introduction

Lignocellulose is the complex matrix of cellulose, hemicellulose, and lignin that constitutes the plant cell wall. The commonly perceived process for lignocellulose use in a biorefinery for ethanol production comprises of two main steps; hydrolysis and fermentation. Hydrolysis refers to the breakdown of polysaccharides present in biomass to monosaccharides and fermentation refers to the use of these monosaccharides for the synthesis of the end-product. Since each set-up has different requirements in terms of physical and chemical conditions, different biomass-degrading enzyme mixtures are required for the efficient release of the available sugars. When hydrolysis and fermentation are performed simultaneously, the temperature is maintained at 30◦C, which is not necessarily the optimum temperature for the commercial enzyme mixtures. When hydrolysis is performed separately from fermentation, the temperature can be closer to the optimum for the enzyme mixture, but other factors—such as product inhibition—can affect the efficiency of the process (Ask et al, 2012)

Methods

Results

Conclusion