Manipulation of malic enzyme in Saccharomyces cerevisiae for increasing NADPH production capacity aerobically in different cellular compartments

Santos, Margarida Moreira dos; Raghevendran, Vijayendran; Kötter, Peter; Olsson, Lisbeth; Nielsen, Jens

doi:10.1016/j.ymben.2004.06.002

Cited by 80 publications

(62 citation statements)

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“…A previous effort to reduce overflow metabolism in S. cerevisiae by manipulating redox balance included deleting GDH1 (encoding cytosolic NADPH-dependent glutamate dehydrogenase), which slightly reduced glycerol formation (17). The combined overexpression of malic enzyme and pyruvate carboxylase resulted in increased NADPH formation at the expense of NADH and ATP formation, but there was no effect on overflow metabolism (18). These results suggest that NADP(H) has a minor role in controlling overflow metabolism in S. cerevisiae.…”

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confidence: 89%

Increasing NADH oxidation reduces overflow metabolism in Saccharomyces cerevisiae

Vemuri

Eiteman

McEwen

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

321

273

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Respiratory metabolism plays an important role in energy production in the form of ATP in all aerobically growing cells. However, a limitation in respiratory capacity results in overflow metabolism, leading to the formation of byproducts, a phenomenon known as ''overflow metabolism'' or ''the Crabtree effect.'' The yeast Saccharomyces cerevisiae has served as an important model organism for studying the Crabtree effect. When subjected to increasing glycolytic fluxes under aerobic conditions, there is a threshold value of the glucose uptake rate at which the metabolism shifts from purely respiratory to mixed respiratory and fermentative. It is well known that glucose repression of respiratory pathways occurs at high glycolytic fluxes, resulting in a decrease in respiratory capacity. Despite many years of detailed studies on this subject, it is not known whether the onset of the Crabtree effect is due to limited respiratory capacity or is caused by glucose-mediated repression of respiration. When respiration in S. cerevisiae was increased by introducing a heterologous alternative oxidase, we observed reduced aerobic ethanol formation. In contrast, increasing nonrespiratory NADH oxidation by overexpression of a water-forming NADH oxidase reduced aerobic glycerol formation. The metabolic response to elevated alternative oxidase occurred predominantly in the mitochondria, whereas NADH oxidase affected genes that catalyze cytosolic reactions. Moreover, NADH oxidase restored the deficiency of cytosolic NADH dehydrogenases in S. cerevisiae. These results indicate that NADH oxidase localizes in the cytosol, whereas alternative oxidase is directed to the mitochondria.alternative oxidase ͉ Crabtree effect ͉ NADH oxidase ͉ redox metabolism R edox homeostasis is a fundamental requirement for sustained metabolism and growth in all biological systems. The intracellular redox potential is primarily determined by the NADH/NAD ratio and to a lesser extent by the NADPH/NADP ratio. In Saccharomyces cerevisiae, Ͼ200 reactions involve these cofactors spread over a large spectrum of cellular functions (1). Because NADH is a highly connected metabolite in the metabolic network (1), any change in the NADH/NAD ratio leads to widespread changes in metabolism (2). NADH is generated primarily in the cytosol by glycolysis and in the mitochondria by the tricarboxylic acid (TCA) cycle. Because the NADH/NAD redox couple cannot traverse the mitochondrial membrane in S. cerevisiae and other eukaryotic cells (3), distinct mechanisms oxidize NADH to NAD in the cytosol and mitochondria. Cytosolic NADH is oxidized by two external (cytosolic) mitochondrial membrane-bound NADH dehydrogenases encoded by NDE1 and NDE2 genes with catalytic sites facing the cytosol (4). Additionally, glycerol-3-phosphate dehydrogenases (encoded by GPD1 and GPD2) oxidize cytosolic NADH with concomitant glycerol formation when the NADH formation rate surpasses its oxidation rate (5). Mitochondrial NADH is oxidized by one internal mitochondrial membrane-bound NADH dehydrogena...

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confidence: 89%

Increasing NADH oxidation reduces overflow metabolism in Saccharomyces cerevisiae

Vemuri

Eiteman

McEwen

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

321

273

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“…to improve specific biotechnological applications. However, to our knowledge, no prokaryotic ME has been used to increase intracellular NADPH levels, as has been done in the yeast Saccharomyces cerevisiae [285,286]. Instead, prokaryotic MEs have been overexpressed for the formation of C 4 -dicarboxylic acids from glucose or pyruvate [287][288][289][290].…”

Section: Malic Enzymementioning

confidence: 99%

Thermococcus kodakarensis : the key to affordable biohydrogen production

Spaans¹

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“…63 Overexpressing ME has been demonstrated as an effective method to enhance lipid accumulation in some strains, such as Mucor circinelloides 59 and S. cerevisiae. 50 Thioesterases is responsible for the release of fatty acid chains from the acyl carrier protein and the production of free fatty acids. Overexpression of thioesterases is also able to increase fatty acid production substantially in many strains such as E. coli and cyanobacteria.…”

Section: Genetic Engineering In Fatty Acids Pathwaymentioning

confidence: 99%

“…During the fatty acids biosynthesis, NADPH is one of the important substrates required for two reduction steps in the fatty acid elongation cycle, which is mainly produced by malic enzyme (ME) and the pentose phosphate pathway. 50 Fatty acid produced in the cell will then be transformed to various fatty acid esters or be degraded. β-oxidation is the principal metabolic pathway responsible for the degradation of fatty acids, which is taking place in the peroxisome, 51 although the transport mechanism remains unclear.…”

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confidence: 99%