Abstract
Bioethanol production processes with Saccharomyces cerevisiae using lignocellulosic biomass as feedstock are challenged by the simultaneous utilization of pentose and hexose sugars from biomass hydrolysates. The pentose uptake into the cell represents a crucial role for the efficiency of the process. The focus of the here presented study was to understand the uptake and conversion of the pentose l-arabinose in S. cerevisiae and reveal its regulation by d-glucose and d-galactose. Gal2p—the most prominent transporter enabling l-arabinose uptake in S. cerevisiae wild-type strains—has an affinity for the transport of l-arabinose, d-glucose, and d-galactose. d-Galactose was reported for being mandatory for inducing GAL2 expression. GAL2 expression is also known to be regulated by d-glucose-mediated carbon catabolite repression, as well as catabolite inactivation. The results of the present study demonstrate that l-arabinose can be used as sole carbon and energy source by the recombinant industrial strain S. cerevisiae DS61180. RT-qPCR and RNA-Seq experiments confirmed that l-arabinose can trigger its own uptake via the induction of GAL2 expression. Expression levels of GAL2 during growth on l-arabinose reached up to 21% of those obtained with d-galactose as sole carbon and energy source. l-Arabinose-induced GAL2 expression was also subject to catabolite repression by d-glucose. Kinetic investigations of substrate uptake, biomass, and product formation during growth on a mixture of d-glucose/l-arabinose revealed impairment of growth and ethanol production from l-arabinose upon d-glucose depletion. The presence of d-glucose is thus preventing the fermentation of l-arabinose in S. cerevisiae DS61180. Comparative transcriptome studies including the wild-type and a precursor strain delivered hints for an increased demand in ATP production and cofactor regeneration during growth of S. cerevisiae DS61180 on l-arabinose. Our results thus emphasize that cofactor and energy metabolism demand attention if the combined conversion of hexose and pentose sugars is intended, for example in biorefineries using lignocellulosics.
Highlights
The limited availability of crude oil and the increasing need for the reduction of greenhouse gases has resulted in an increased demand for second generation biofuels as more environmentally friendly energy carriers [1,2,3,4]
Despite it is frequently reported that d-galactose is crucial for GAL2 expression in wild-type S. cerevisiae, the present study demonstrates that S. cerevisiae DS61180 is able to induce GAL2 expression by l-arabinose
A metabolic burden caused by strain development is not evident during cultivations on d‐glucose as sole carbon source under aerobic conditions in minimal medium Ethanol production for the industrial strain S. cerevisiae DS61180 has already been reported for a substrate mixture of d-glucose, d-galactose, d-xylose, and l-arabinose [84]
Summary
The limited availability of crude oil and the increasing need for the reduction of greenhouse gases has resulted in an increased demand for second generation biofuels as more environmentally friendly energy carriers [1,2,3,4]. For the handling of mixed sugar fermentations and the development of flexible production strains, it is important to understand how the single components of the system affect the physiology of the fermenting biocatalyst. This holds especially true for studying the effects of the respective sugars types and its combinations on the productivity of whole-cell biocatalysts with a complex substrate-dependent metabolism like S. cerevisiae, which is regulated by the availability of carbon sources via, e.g., carbon catabolite repression [13]. An insufficient understanding of the genetic background can impair strain improvement [14]
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