Saccharomyces cerevisiae phosphoglucose isomerase and fructose bisphosphate aldolase can be replaced functionally by the corresponding enzymes of Escherichia coli and Drosophila melanogaster

Boles, Eckhard; Zimmermann, Friedrich K.

doi:10.1007/bf00351494

Cited by 38 publications

(32 citation statements)

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“…4; Cabib and Leloir, 1958;Vandercammen et al, 1989;Londesborough and Vuorio, 1991). However, even at only 1 mM phosphate, the concentration of Fru6P for half-maximal activation (1.4 mM), is higher than the bulk concentration of Fru6P in yeast ( 5 0.1 -0.7 mM; Lagunas and Gancedo, 1983;Boles and Zimmerman, 1993). FranSois et al (1991) reported that the sum of Glc6P and Fru6P was only 0.4 mM during trehalose accumulation.…”

Section: Discussionmentioning

confidence: 99%

“…In this region, Glc6P and Fru6P concentrations may differ radically from their bulk cytosolic values. Finally, it is curious that the phenotype of yeast lacking phosphoglucoisomerase resembles that of cifl andfdpl mutants: both are characterized by poor growth in the presence of glucose and defective glucose-induced cAMP signalling, which cannot be completely explained by their over-accumulation of Glc6P and depletion of ATP (Bafiuelos and Fraenkel, 1982;Boles and Zimmerman, 1993). Possibly an appropriate Glc6PIFru6P ratio is required for the correct functioning of the 56-kDa polypeptide also in its role as a component of the cAMP signalling system.…”

Section: Discussionmentioning

confidence: 99%

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Purification of trehalose synthase from baker's yeast

Londesborough¹,

Vuorio²

1993

European Journal of Biochemistry

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A trehalose synthase purified from baker's yeast contained 56-kDa, 102-kDa and 123-kDa polypeptides as its main components. The 102-kDa polypeptide was isolated and shown to be a specific trehalose-6-phosphatase. The trehalose-6-phosphate synthase (Tre6P synthase) activator described by Londesborough and Vuorio [(1991) J. Gen. Microbiol. 137,323-3301 was shown to be phosphoglucoisomerase and to function entirely by generating fructose 6-phosphate. Below 35 "C, fructose 6-phosphate is a powerful activator of the Tre6P synthase activity of intact trehalose synthase, especially at physiological phosphate concentration, but does not affect its trehalose-6-phosphatase activity nor the Tre6P synthase activity of truncated trehalose synthase containing truncated versions of the 123-kDa polypeptide. At 50"C, activation by fructose 6-phosphate and inhibition by phosphate are greatly decreased, resulting in an unusually high temperature-dependence for the Tre6P synthase activity at a physiological phosphate concentration (2 mM).Recent findings, summarized by Wiemken (1990), suggest that the primary role of trehalose in yeast is as a protectant against a variety of stresses, including extremes of temperature and desiccation. Cabib and Leloir (1958) showed that trehalose is synthesized in yeast from glucose 6-phosphate (Glc6P) and uridinediphosphoglucose (UDP-Glc) via the intermediate trehalose 6-phosphate (Tre6P). Londesborough and Vuorio (1991) purified a yeast protein that exhibited both Tre6P synthase and Tre6P phosphatase activities and contained a short polypeptide chain (about 57 kDa, now known to be 56.3 kDa) and two fragments (86 and 93 kDa) apparently derived from a single long chain. We shall now call this preparation truncated trehalose synthase.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Purification of trehalose synthase from baker's yeast

Londesborough¹,

Vuorio²

1993

European Journal of Biochemistry

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“…Molecular techniques were performed according to published procedures (22). Yeast transformations and reisolation of plasmid DNA from yeast cells were carried out as described previously (6,11).…”

Section: Methodsmentioning

confidence: 99%

Codon-Optimized Bacterial Genes Improve l -Arabinose Fermentation in Recombinant Saccharomyces cerevisiae

Wiedemann

Boles

2008

Appl Environ Microbiol

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108

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Bioethanol produced by microbial fermentations of plant biomass hydrolysates consisting of hexose and pentose mixtures is an excellent alternative to fossil transportation fuels. However, the yeast Saccharomyces cerevisiae, commonly used in bioethanol production, can utilize pentose sugars like L-arabinose or D-xylose only after heterologous expression of corresponding metabolic pathways from other organisms. Here we report the improvement of a bacterial L-arabinose utilization pathway consisting of L-arabinose isomerase from Bacillus subtilis and L-ribulokinase and L-ribulose-5-P 4-epimerase from Escherichia coli after expression of the corresponding genes in S. cerevisiae. L-Arabinose isomerase from B. subtilis turned out to be the limiting step for growth on L-arabinose as the sole carbon source. The corresponding enzyme could be effectively replaced by the enzyme from Bacillus licheniformis, leading to a considerably decreased lag phase. Subsequently, the codon usage of all the genes involved in the L-arabinose pathway was adapted to that of the highly expressed genes encoding glycolytic enzymes in S. cerevisiae. Yeast transformants expressing the codon-optimized genes showed strongly improved L-arabinose conversion rates. With this rational approach, the ethanol production rate from L-arabinose could be increased more than 2.5-fold from 0.014 g ethanol h ؊1 (g dry weight) Decreasing fossil energy resources and the climate change caused by emissions of CO 2 from their burning have led to a growing interest in renewable-energy alternatives. Bioethanol produced by microbial fermentations of plant biomass is an excellent alternative to fossil fuels. However, for economical ethanol production, it is not enough to convert only the starch and sucrose fractions of plant biomass as is mainly done in conventional ethanol plants. The importance of the conversion of the lignocellulosic fraction as well becomes more and more evident. Lignocellulosic hydrolysates consist of easily fermentable hexose sugars but also significant amounts of pentose sugars. Depending on the choice of raw material, the amounts of pentoses found in lignocellulosic hydrolysates range from, for example, 16% xylan and 5% arabinan in grass and 15% arabinan and 19% xylan in wheat bran (14). These numbers clearly indicate that both hexoses and pentoses must be fermented in an efficient ethanol production process. Even small increases in substrate utilization should significantly improve the overall process costs (28).Although Saccharomyces cerevisiae is the organism most widely used for ethanol production and is able to convert hexoses rapidly and with high ethanol yields, wild-type S. cerevisiae strains are not able to ferment pentose sugars, such as D-xylose and L-arabinose, efficiently. Even though xylose can be slowly metabolized, at least by adapted strains (2), the second most abundant pentose, L-arabinose, cannot be converted at all. Several different genetic-engineering approaches have been used in attempts to enable D-xylose and L-arabinose...

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“…Molecular biology techniques were performed by using published procedures (25). Yeast transformations and reisolation of plasmid DNA from yeast cells were carried out as described previously (3,12).…”

Section: Methodsmentioning

confidence: 99%

A Modified Saccharomyces cerevisiae Strain That Consumes l -Arabinose and Produces Ethanol

Becker

Boles

2003

Appl Environ Microbiol

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234

208

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Metabolic engineering is a powerful method to improve, redirect, or generate new metabolic reactions or whole pathways in microorganisms. Here we describe the engineering of a Saccharomyces cerevisiae strain able to utilize the pentose sugar L-arabinose for growth and to ferment it to ethanol. Expanding the substrate fermentation range of S. cerevisiae to include pentoses is important for the utilization of this yeast in economically feasible biomass-to-ethanol fermentation processes. After overexpression of a bacterial L-arabinose utilization pathway consisting of Bacillus subtilis AraA and Escherichia coli AraB and AraD and simultaneous overexpression of the L-arabinose-transporting yeast galactose permease, we were able to select an L-arabinose-utilizing yeast strain by sequential transfer in L-arabinose media. Molecular analysis of this strain, including DNA microarrays, revealed that the crucial prerequisite for efficient utilization of L-arabinose is a lowered activity of L-ribulokinase. Moreover, high L-arabinose uptake rates and enhanced transaldolase activities favor utilization of L-arabinose. With a doubling time of about 7.9 h in a medium with L-arabinose as the sole carbon source, an ethanol production rate of 0.06 to 0.08 g of ethanol per g (dry weight) ⅐ h ؊1 under oxygen-limiting conditions, and high ethanol yields, this yeast strain should be useful for efficient fermentation of hexoses and pentoses in cellulosic biomass hydrolysates.

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Saccharomyces cerevisiae phosphoglucose isomerase and fructose bisphosphate aldolase can be replaced functionally by the corresponding enzymes of Escherichia coli and Drosophila melanogaster

Cited by 38 publications

References 39 publications

Purification of trehalose synthase from baker's yeast

Purification of trehalose synthase from baker's yeast

Codon-Optimized Bacterial Genes Improve l -Arabinose Fermentation in Recombinant Saccharomyces cerevisiae

A Modified Saccharomyces cerevisiae Strain That Consumes l -Arabinose and Produces Ethanol

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