Abstract BackgroundCost-effective production of biofuels from lignocellulose requires the fermentation ofd-xylose. Many yeast species within and closely related to the generaSpathasporaandScheffersomyces(both of the order Serinales) natively assimilate and ferment xylose. Other species consume xylose inefficiently, leading to extracellular accumulation of xylitol. Xylitol excretion is thought to be due to the different cofactor requirements of the first two steps of xylose metabolism. Xylose reductase (XR) generally uses NADPH to reduce xylose to xylitol, while xylitol dehydrogenase (XDH) generally uses NAD+to oxidize xylitol to xylulose, creating an imbalanced redox pathway. This imbalance is thought to be particularly consequential in hypoxic or anoxic environments. ResultsWe screened the growth of xylose-fermenting yeast species in high and moderate aeration and identified both ethanol producers and xylitol producers. Selected species were further characterized for their XR and XDH cofactor preferences by enzyme assays and gene expression patterns by RNA-Seq. Our data revealed that xylose metabolism is more redox balanced in some species, but it is strongly affected by oxygen levels. Under high aeration, most species switched from ethanol production to xylitol accumulation, despite the availability of ample oxygen to accept electrons from NADH. This switch was followed by decreases in enzyme activity and the expression of genes related to xylose metabolism, suggesting that bottlenecks in xylose fermentation are not always due to cofactor preferences. Finally, we expressedXYLgenes from multipleScheffersomycesspecies in a strain ofSaccharomyces cerevisiae. RecombinantS. cerevisiaeexpressingXYL1fromScheffersomyces xylosifermentans, which encodes an XR without a cofactor preference, showed improved anaerobic growth on xylose as the primary carbon source compared toS. cerevisiaestrain expressingXYLgenes fromScheffersomyces stipitis. ConclusionCollectively, our data do not support the hypothesis that xylitol accumulation occurs primarily due to differences in cofactor preferences between xylose reductase and xylitol dehydrogenase; instead, gene expression plays a major role in response to oxygen levels. We have also identified the yeastSc. xylosifermentansas a potential source for genes that can be engineered intoS. cerevisiaeto improve xylose fermentation and biofuel production.
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Codon Optimization Improves the Prediction of Xylose Metabolism from Gene Content in Budding Yeasts
Abstract Xylose is the second most abundant monomeric sugar in plant biomass. Consequently, xylose catabolism is an ecologically important trait for saprotrophic organisms, as well as a fundamentally important trait for industries that hope to convert plant mass to renewable fuels and other bioproducts using microbial metabolism. Although common across fungi, xylose catabolism is rare within Saccharomycotina, the subphylum that contains most industrially relevant fermentative yeast species. The genomes of several yeasts unable to consume xylose have been previously reported to contain the full set of genes in the XYL pathway, suggesting the absence of a gene–trait correlation for xylose metabolism. Here, we measured growth on xylose and systematically identified XYL pathway orthologs across the genomes of 332 budding yeast species. Although the XYL pathway coevolved with xylose metabolism, we found that pathway presence only predicted xylose catabolism about half of the time, demonstrating that a complete XYL pathway is necessary, but not sufficient, for xylose catabolism. We also found that XYL1 copy number was positively correlated, after phylogenetic correction, with xylose utilization. We then quantified codon usage bias of XYL genes and found that XYL3 codon optimization was significantly higher, after phylogenetic correction, in species able to consume xylose. Finally, we showed that codon optimization of XYL2 was positively correlated, after phylogenetic correction, with growth rates in xylose medium. We conclude that gene content alone is a weak predictor of xylose metabolism and that using codon optimization enhances the prediction of xylose metabolism from yeast genome sequence data.
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- PAR ID:
- 10426545
- Editor(s):
- Townsend, Jeffrey
- Date Published:
- Journal Name:
- Molecular Biology and Evolution
- Volume:
- 40
- Issue:
- 6
- ISSN:
- 0737-4038
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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