Browsing by Author "Kruger, Francois"

Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    Engineering saccharomyces cerevisiae for application in integrated bioprocessing biorefineries
    (Elsevier Ltd, 2024) Minnaar, Letitia S; Kruger, Francois; Fortuin, Jordan
    After decades of research and development, no organism — natural or engineered — has been described that can produce commodity products through direct microbial conversion to meet industry demands in terms of rates and yields. Variation in lignocellulosic biomass (LCB) feedstocks, the lack of a widely applicable pretreatment method, and the limited economic value of energy products further complicates second-generation biofuel production. Nevertheless, the emergence of advanced genomic editing tools and a more comprehensive understanding of yeast metabolic systems offer promising avenues for the creation of yeast strains tailored to LCB biorefineries. Here, we discuss recent advances toward developing yeast strains that could convert different LCB fractions into a series of economically viable commodity products in a biorefinery.
  • Thumbnail Image
    Item
    Engineering xylan assimilation into industrial strains of Saccharomyces cerevisiae.
    (University of the Western Cape, 2025) Kruger, Francois
    Second-generation biofuels are attractive alternatives to environmentally damaging, non-renewable fossil fuels, as they are carbon neutral and produced from renewable lignocellulosic biomass (LCB). One of the main challenges facing LCB conversion to bioethanol is the incomplete use of all available sugars present in the biomass. To overcome this challenge, the hemicellulose fraction, consisting mostly of xylan, should be targeted for conversion in addition to the cellulose fraction. This study aimed to address this issue by developing strains of Saccharomyces cerevisiae capable of xylan degradation and efficient utilization of xylose. The laboratory strain S288C was the initial candidate used. The strain was previously engineered with a xylose isomerase (XI)pathway and subsequently further engineered using CRISPR-Cas9 technology. GH43xylosidase activity, either freely secreted or tethered to the cells, was introduced into the yeast, together with secreted xylanase activity. The strains were evaluated through enzymatic assays, growth on media containing xylo-oligosaccharides (XOS) and xylan, and fermentations on xylan media. It was found that the strain with tethered xylosidase and secreted xylanase activity showed the best growth on the polymeric substrates (XOS or xylan) and produced the highest ethanol titre of 0.47 g/L during fermentation on xylan as carbon source. Natural S. cerevisiae strain isolates YI13, YI59 and FIN1 were selected for potential industrial applications due to their robust fermentation performance and enhanced ethanol production compared to the reference strain S288C. Efficient xylose utilization was conferred to the natural strains through engineering with an XI gene cassette and a xylose transporter, combined with adaptive laboratory evolution (ALE) in minimal media with xylose as the sole carbon source. The xylose-utilizing strains were further engineered with cell-associated GH43 xylosidase activity and secreted xylanase activity. Enzymatic assays, growth trials on hemicellulosic substrates and fermentation on xylose and xylan showed that the strains were successfully engineered with xylan conversion capabilities and efficient xylose utilization. The final engineered version of YI13 showed the best xylose and xylan conversion, with ethanol titres of 4.51 g/L from xylose and ~ 3 g/L from xylan. This is the highest reported level of ethanol produced from polymeric xylan to date.