Abstract
We have recently demonstrated that heterologous expression of a bacterial xylose isomerase gene (xylA) of Burkholderia cenocepacia enabled a laboratorial Saccharomyces cerevisiae strain to ferment xylose anaerobically, without xylitol accumulation. However, the recombinant yeast fermented xylose slowly. In this study, an evolutionary engineering strategy was applied to improve xylose fermentation by the xylA-expressing yeast strain, which involved sequential batch cultivation on xylose. The resulting yeast strain co-fermented glucose and xylose rapidly and almost simultaneously, exhibiting improved ethanol production and productivity. It was also observed that when cells were grown in a medium containing higher glucose concentrations before being transferred to fermentation medium, higher rates of xylose consumption and ethanol production were obtained, demonstrating that xylose utilization was not regulated by catabolic repression. Results obtained by qPCR demonstrate that the efficiency in xylose fermentation showed by the evolved strain is associated, to the increase in the expression of genes HXT2 and TAL1, which code for a low-affinity hexose transporter and transaldolase, respectively. The ethanol productivity obtained after the introduction of only one genetic modification and the submission to a one-stage process of evolutionary engineering was equivalent to those of strains submitted to extensive metabolic and evolutionary engineering, providing solid basis for future applications of this strategy in industrial strains.Electronic supplementary materialThe online version of this article (doi:10.1186/s13568-015-0102-y) contains supplementary material, which is available to authorized users.
Highlights
The ethanol production from lignocellulosic biomass is a promising alternative energy source
We report an evolutionary engineering approach which promoted an increase on the consumption rate of xylose and ethanol production by the S. cerevisiae expressing xylose isomerase from B. cenocepacia
Evolutionary engineering promotes high levels of xylose consumption in the presence of glucose The adaptive evolution experiments were performed with 40 passages in YNB medium and isolated clones were screened
Summary
The ethanol production from lignocellulosic biomass is a promising alternative energy source. A great diversity of metabolic engineering and adaptation approaches have been tested to boost xylose fermentation in biomass hydrolysates, yield and productivity of ethanol by genetically engineered S. cerevisiae strains are still much lower than those of glucose fermentation. Discovery or engineering of a xylosespecific transporter which is not inhibited by glucose and which shows high affinity and capacity of transport might improve cellular performance to ferment xylose in biomass hydrolysates (Weber et al 2010). We described the functional expression of Burkholderia cenocepacia xylose isomerase and its effect on the ability of S. cerevisiae to ferment xylose-glucose blends (De Figueiredo Vilela et al 2013). We report an evolutionary engineering approach which promoted an increase on the consumption rate of xylose and ethanol production by the S. cerevisiae expressing xylose isomerase from B. cenocepacia. Aiming to unravel the changes between the metabolically engineered and the subsequently evolved strain, the expression profile of several genes involved in xylose fermentation was analyzed
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