Overexpression of NADH-dependent fumarate reductase improves D-xylose fermentation in recombinant Saccharomyces cerevisiae. 2013

Laura Salusjärvi, and Sanna Kaunisto, and Sami Holmström, and Maija-Leena Vehkomäki, and Kari Koivuranta, and Juha-Pekka Pitkänen, and Laura Ruohonen
VTT, Technical Research Centre of Finland, PO Box 1000, 02044, VTT, Finland, laura.salusjarvi@vtt.fi.

Deviation from optimal levels and ratios of redox cofactors NAD(H) and NADP(H) is common when microbes are metabolically engineered. The resulting redox imbalance often reduces the rate of substrate utilization as well as biomass and product formation. An example is the metabolism of D-xylose by recombinant Saccharomyces cerevisiae strains expressing xylose reductase and xylitol dehydrogenase encoding genes from Scheffersomyces stipitis. This pathway requires both NADPH and NAD(+). The effect of overexpressing the glycosomal NADH-dependent fumarate reductase (FRD) of Trypanosoma brucei in D-xylose-utilizing S. cerevisiae alone and together with an endogenous, cytosol directed NADH-kinase (POS5Δ17) was studied as one possible solution to overcome this imbalance. Expression of FRD and FRD + POS5Δ17 resulted in 60 and 23 % increase in ethanol yield, respectively, on D-xylose under anaerobic conditions. At the same time, xylitol yield decreased in the FRD strain suggesting an improvement in redox balance. We show that fumarate reductase of T. brucei can provide an important source of NAD(+) in yeast under anaerobic conditions, and can be useful for metabolic engineering strategies where the redox cofactors need to be balanced. The effects of FRD and NADH-kinase on aerobic and anaerobic D-xylose and D-glucose metabolism are discussed.

UI MeSH Term Description Entries
D009243 NAD A coenzyme composed of ribosylnicotinamide 5'-diphosphate coupled to adenosine 5'-phosphate by pyrophosphate linkage. It is found widely in nature and is involved in numerous enzymatic reactions in which it serves as an electron carrier by being alternately oxidized (NAD+) and reduced (NADH). (Dorland, 27th ed) Coenzyme I,DPN,Diphosphopyridine Nucleotide,Nadide,Nicotinamide-Adenine Dinucleotide,Dihydronicotinamide Adenine Dinucleotide,NADH,Adenine Dinucleotide, Dihydronicotinamide,Dinucleotide, Dihydronicotinamide Adenine,Dinucleotide, Nicotinamide-Adenine,Nicotinamide Adenine Dinucleotide,Nucleotide, Diphosphopyridine
D010100 Oxygen An element with atomic symbol O, atomic number 8, and atomic weight [15.99903; 15.99977]. It is the most abundant element on earth and essential for respiration. Dioxygen,Oxygen-16,Oxygen 16
D011994 Recombinant Proteins Proteins prepared by recombinant DNA technology. Biosynthetic Protein,Biosynthetic Proteins,DNA Recombinant Proteins,Recombinant Protein,Proteins, Biosynthetic,Proteins, Recombinant DNA,DNA Proteins, Recombinant,Protein, Biosynthetic,Protein, Recombinant,Proteins, DNA Recombinant,Proteins, Recombinant,Recombinant DNA Proteins,Recombinant Proteins, DNA
D005285 Fermentation Anaerobic degradation of GLUCOSE or other organic nutrients to gain energy in the form of ATP. End products vary depending on organisms, substrates, and enzymatic pathways. Common fermentation products include ETHANOL and LACTIC ACID. Fermentations
D005947 Glucose A primary source of energy for living organisms. It is naturally occurring and is found in fruits and other parts of plants in its free state. It is used therapeutically in fluid and nutrient replacement. Dextrose,Anhydrous Dextrose,D-Glucose,Glucose Monohydrate,Glucose, (DL)-Isomer,Glucose, (alpha-D)-Isomer,Glucose, (beta-D)-Isomer,D Glucose,Dextrose, Anhydrous,Monohydrate, Glucose
D012441 Saccharomyces cerevisiae A species of the genus SACCHAROMYCES, family Saccharomycetaceae, order Saccharomycetales, known as "baker's" or "brewer's" yeast. The dried form is used as a dietary supplement. Baker's Yeast,Brewer's Yeast,Candida robusta,S. cerevisiae,Saccharomyces capensis,Saccharomyces italicus,Saccharomyces oviformis,Saccharomyces uvarum var. melibiosus,Yeast, Baker's,Yeast, Brewer's,Baker Yeast,S cerevisiae,Baker's Yeasts,Yeast, Baker
D014346 Trypanosoma brucei brucei A hemoflagellate subspecies of parasitic protozoa that causes nagana in domestic and game animals in Africa. It apparently does not infect humans. It is transmitted by bites of tsetse flies (Glossina). Trypanosoma brucei,Trypanosoma brucei bruceus,Trypanosoma bruceus,brucei brucei, Trypanosoma,brucei, Trypanosoma brucei,bruceus, Trypanosoma,bruceus, Trypanosoma brucei
D014993 Xylitol A five-carbon sugar alcohol derived from XYLOSE by reduction of the carbonyl group. It is as sweet as sucrose and used as a noncariogenic sweetener.
D014994 Xylose D-Xylose,D Xylose
D044925 Oxidoreductases Acting on CH-CH Group Donors A subclass of enzymes which includes all dehydrogenases acting on carbon-carbon bonds. This enzyme group includes all the enzymes that introduce double bonds into substrates by direct dehydrogenation of carbon-carbon single bonds. Oxidoreductases Acting on CH CH Group Donors

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