Search

Abstract Background The need to restructure the world’s energy matrix based on fossil fuels and mitigate greenhouse gas emissions stimulated the development of new biobased technologies for renewable energy. One promising and cleaner alternative is the use of second-generation (2G) fuels, produced from lignocellulosic biomass sugars. A major challenge on 2G technologies establishment is the inefficient assimilation of the five-carbon sugar xylose by engineered Saccharomyces cerevisiae strains, increasing fermentation time. The uptake of xylose across the plasma membrane is a critical limiting step and the budding yeast S. cerevisiae is not designed with a broad transport system and regulatory mechanisms to assimilate xylose in a wide range of concentrations present in 2G processes. Results Assessing diverse microbiomes such as the digestive tract of plague insects and several decayed lignocellulosic biomasses, we isolated several yeast species capable of using xylose. Comparative fermentations selected the yeast Candida sojae as a potential source of high-affinity transporters. Comparative genomic analysis elects four potential xylose transporters whose properties were evaluated in the transporter null EBY.VW4000 strain carrying the xylose-utilizing pathway integrated into the genome. While the traditional xylose transporter Gxf1 allows an improved growth at lower concentrations (10 g/L), strains containing Cs3894 and Cs4130 show opposite responses with superior xylose uptake at higher concentrations (up to 50 g/L). Docking and normal mode analysis of Cs4130 and Gxf1 variants pointed out important residues related to xylose transport, identifying key differences regarding substrate translocation comparing both transporters. Conclusions Considering that xylose concentrations in second-generation hydrolysates can reach high values in several designed processes, Cs4130 is a promising novel candidate for xylose uptake. Here, we demonstrate a novel eukaryotic molecular transporter protein that improves growth at high xylose concentrations and can be used as a promising target towards engineering efficient pentose utilization in yeast.

Background: The need to restructure the world's energy matrix based on fossil fuels and mitigate greenhouse gas emissions stimulated the development of new biobased technologies for renewable energy. One promising and cleaner alternative is the use of second-generation (2G) fuels, produced from lignocellulosic biomass sugars. A major challenge on 2G technologies establishment is the inefficient assimilation of the five-carbon sugar xylose by engineered Saccharomyces cerevisiae strains, increasing fermentation time. The uptake of xylose across the plasma membrane is a critical limiting step and the budding yeast S. cerevisiae is not designed with a broad transport system and regulatory mechanisms to assimilate xylose in a wide range of concentrations present in 2G processes. Results: Assessing diverse microbiomes such as the digestive tract of plague insects and several decayed lignocellulosic biomasses, we isolated several yeast species capable of using xylose. Comparative fermentations selected the yeast Candida sojae as a potential source of high-affinity transporters. Comparative genomic analysis elects four potential xylose transporters whose properties were evaluated in the transporter null EBY.VW4000 strain carrying the xylose-utilizing pathway integrated into the genome. While the traditional xylose transporter Gxf1 allows an improved growth at lower concentrations (10 g/L), strains containing Cs3894 and Cs4130 show opposite responses with superior xylose uptake at higher concentrations (up to 50 g/L). Docking and normal mode analysis of Cs4130 and Gxf1 variants pointed out important residues related to xylose transport, identifying key differences regarding substrate translocation comparing both transporters. Conclusions: Considering that xylose concentrations in second-generation hydrolysates can reach high values in several designed processes, Cs4130 is a promising novel candidate for xylose uptake. Here, we demonstrate a novel eukaryotic molecular transporter protein that improves growth at high xylose concentrations and can be used as a promising target towards engineering efficient pentose utilization in yeast. [ABSTRACT FROM AUTHOR]

Background: An economically viable production of biofuels and biochemicals from lignocellulose requires microorganisms that can readily convert both the cellulosic and hemicellulosic fractions into product. The yeast Candida intermedia displays a high capacity for uptake and conversion of several lignocellulosic sugars including the abundant pentose d-xylose, an underutilized carbon source since most industrially relevant microorganisms cannot naturally ferment it. Thus, C. intermedia constitutes an important source of knowledge and genetic information that could be transferred to industrial microorganisms such as Saccharomyces cerevisiae to improve their capacity to ferment lignocellulose-derived xylose. Results: To understand the genetic determinants that underlie the metabolic properties of C. intermedia, we sequenced the genomes of both the in-house-isolated strain CBS 141442 and the reference strain PYCC 4715. De novo genome assembly and subsequent analysis revealed C. intermedia to be a haploid species belonging to the CTG clade of ascomycetous yeasts. The two strains have highly similar genome sizes and number of protein-encoding genes, but they differ on the chromosomal level due to numerous translocations of large and small genomic segments. The transcriptional profiles for CBS 141442 grown in medium with either high or low concentrations of glucose and xylose were determined through RNA-sequencing analysis, revealing distinct clusters of co-regulated genes in response to different specific growth rates, carbon sources and osmotic stress. Analysis of the genomic and transcriptomic data also identified multiple xylose reductases, one of which displayed dual NADH/NADPH co-factor specificity that likely plays an important role for co-factor recycling during xylose fermentation. Conclusions: In the present study, we performed the first genomic and transcriptomic analysis of C. intermedia and identified several novel genes for conversion of xylose. Together the results provide insights into the mechanisms underlying saccharide utilization in C. intermedia and reveal potential target genes to aid in xylose fermentation in S. cerevisiae. [ABSTRACT FROM AUTHOR]