Abstract
BackgroundFermentation of lignocellulosic biomass is an attractive alternative for the production of bioethanol. Traditionally, the yeast Saccharomyces cerevisiae is used in industrial ethanol fermentations. However, S. cerevisiae is naturally not able to ferment the pentose sugars D-xylose and L-arabinose, which are present in high amounts in lignocellulosic raw materials.ResultsWe describe the engineering of laboratory and industrial S. cerevisiae strains to co-ferment the pentose sugars D-xylose and L-arabinose. Introduction of a fungal xylose and a bacterial arabinose pathway resulted in strains able to grow on both pentose sugars. Introduction of a xylose pathway into an arabinose-fermenting laboratory strain resulted in nearly complete conversion of arabinose into arabitol due to the L-arabinose reductase activity of the xylose reductase. The industrial strain displayed lower arabitol yield and increased ethanol yield from xylose and arabinose.ConclusionOur work demonstrates simultaneous co-utilization of xylose and arabinose in recombinant strains of S. cerevisiae. In addition, the co-utilization of arabinose together with xylose significantly reduced formation of the by-product xylitol, which contributed to improved ethanol production.
Highlights
Large interest in biofuel ethanol production from renewable sources has led in numerous attempts to construct microbial strains that efficiently ferment lignocellulose hydrolysate to ethanol [1,2]
MATa leu2-3,112 ura3-52 trp1-289 his3-∆1 MAL2-8c SUC2, unknown beneficial mutations for arabinose utilization, with plasmids YEparaA, YEparaBG361A, YEparaD, YEpGAL2 MATa leu2-3,112 ura3-52 trp1-289 his3-∆1 MAL2-8c SUC2, unknown beneficial mutations for arabinose utilization, HIS3::YIpXR/xylitol dehydrogenase (XDH)/XK BWY02.X with plasmids p424H7, p425H7, p426H7 BWY02.X with plasmids YEpURAaraA, YEparaBG361A, YEparaD HIS3::pPGK-Xyl1-tPGK, pADH-Xyl2-tADH, pPGK-XKS1-tPGK TMB 3400, TRP1::YIpAraB KanMX TMB 3060, NTS2::pHXT7-arabinose isomerase (AraA)-tCYC1and NTS2::pHXT7-AraD-tCYC1, KanMX TMB 3061 retransformed with NTS2::pHXT7-AraA-tCYC1 MATa pPGK1-XYL1-tPGK1, pADH1-XYL2-tADH1, pPGK1-XKS1-tPGK1, HIS3 pHXT7-AraA-tCYC1, HIS3 pHXT7-AraA-tCYC, URA3 pHXT7-AraB-tCYC1, TRP1 pHXT7-AraD-tCYC1, LEU2 pHXT7-MCS-tCYC1, HIS3 pHXT7-MCS-tCYC1, TRP1 pHXT7-MCS-tCYC1, LEU2 pHXT7-MCS-tCYC1, URA3 Cloning vector KanMX, pHXT7-AraB-tCYC1 TRP1 pBluescript, NTS2 prDNA, NTS2::pHXT7-AraA-tCYC1 prDNA, NTS2::pHXT7-AraD-tCYC1
The construct was based on the arabinose-fermenting strain JBY25-4M [11] (Table 1), which contains the bacterial arabinose pathway consisting of the genes for L-arabinose isomerase (AraA), Lribulokinase (AraB) and L-ribulose-5-P 4-epimerase (AraD) as well as the overexpressed endogenous galactose transporter gene (GAL2), each on a separate multicopy plasmid, and which has beneficial mutations resulting from prolonged culturing in arabinose medium [11]
Summary
Large interest in biofuel ethanol production from renewable sources has led in numerous attempts to construct microbial strains that efficiently ferment lignocellulose hydrolysate to ethanol [1,2]. The optimal micro-organism must be robust, ethanol and inhibitor tolerant, viable at low pH, and produce high ethanol yields from all sugars present in lignocellulose [3]. The baker's yeast Saccharomyces cerevisiae is very suitable for industrial ethanol production, it cannot utilize pentoses that represent a significant fraction of the sugars present in lignocellulosic raw materials. The yeast Saccharomyces cerevisiae is used in industrial ethanol fermentations. S. cerevisiae is naturally not able to ferment the pentose sugars D-xylose and L-arabinose, which are present in high amounts in lignocellulosic raw materials
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