Direct Production of Difructose Anhydride IV from Sucrose by Co-fermentation of Recombinant Yeasts

A functional sweetener, difructose anhydride IV (DFA IV), is enzymatically produced from sucrose via levan by levansucrase (LSRase) followed by levan fructotransferase (LFTase). Here, we have demonstrated a consolidated production system for the direct conversion of DFA IV from sucrose using the co-culture of two recombinant yeast strains secreting LSRase from Bacillus subtilis and LFTase from Arthrobacter ureafaciens, respectively. To ensure secretory production of the enzymes, target protein-specific translational fusion partners (TFP) were employed, and the selected strains produced 3.8 U/mL of LSRase and 16.0 U/mL LFTase activity into the fermentation broth. To optimise the direct production, sucrose concentration and cell ratios were investigated. In the optimised conditions, 64.3 g/L crude DFA IV was directly produced from 244.7 g/L sucrose using co-fermentation of recombinant yeasts. These results promise an efficient production titre, yield, and DFA IV productivity in an industrially applicable method.

New Approach of Highly Efficient Fermentation Process for Bioethanol using Xylose as Agriculture Residues

Agricultural waste biomass has already been transferred to bioethanol and used as energy related products, although many issues such as efficiency and productivity still exist to be overcome. In this study, the protein engineering was applied to generate enzymes with completely reversed coenzyme specificity and developed recombinant yeasts containing those engineered enzymes for construction of an efficient biomass-ethanol conversion system. Recombinant yeasts were constructed with the genes encoding a wild type xylose reductase (XR) and the protein engineered xylitol dehydrogenase (XDH) (with NADP) of Pichia stipitis. These recombinant yeasts were characterized based on the enzyme activity and fermentation ability of xylose to ethanol. The protein engineered enzymes were expressed significantly in Saccharomyces cerevisiae as judged by the enzyme activity in vitro. Ethanol fermentation was measured in batch culture under anaerobic conditions. The significant enhancement was found in Y-ARS strain, in which NADP+-dependent XDH was expressed; 85% decrease of unfavorable xylitol excretion with 26% increased ethanol production, when compared with the reference strain expressing the wild–type XDH.

New Approach of Highly Efficient Fermentation Process for Bio ethanol using Xylose as Agricultur e Residues

Agricultural waste biomasshas already been transferred to bioethanol and used as energy related products, although many issues such as efficiency and productivity still exist to be overcome. In this study, the protein engineering was applied to generate enzymes with completely reversed coenzyme specificity and developed recombinant yeasts containing those engineered enzymes for construction of an efficient biomass-ethanol conversion system. Recombinant yeasts were constructed with the genes encoding a wild type xylose reductase (XR)and the protein engineered xylitol dehydrogenase (XDH)(with NADP) of Pichiastipitis. These recombinant yeasts were characterized based on the enzyme activity and fermentation ability of xylose to ethanol. The protein engineered enzymes were expressed significantly in Saccharomycescerevisiaeas judged by the enzyme activity in vitro. Ethanol fermentation was measured in batch culture under anaerobic conditions. The significant enhancement was found in Y-ARSstrain, in which NADP+-dependentXDH was expressed; 85% decrease of unfavorable xylitol excretion with 26% increased ethanol production, when compared with the reference strain expressing the wild–type XDH.