Peculiarities of the Regulation of Fermentation and Respiration in the Crabtree-Negative, Xylose-Fermenting Yeast Pichia stipitis

Passoth, Volkmar; Zimmermann, Martín; Klinner, Ulrich

doi:10.1007/978-1-4612-0223-3_18

Cited by 36 publications

(66 citation statements)

References 28 publications

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“…2), as was also reported for other yeasts (29). This flux ratio is (53), and predominantly respirative in Crabtree-negative yeasts such as P. stipitis (33). Second, during respiro-fermentative metabolism of S. cerevisiae in batch culture, cytosolic OAA originates exclusively from PYR and is transported unidirectionally into the mitochondria (27).…”

Section: Discussionsupporting

confidence: 74%

“…In Crabtree-positive yeasts, such as S. cerevisiae, elevated glucose concentrations induce the carbon catabolite repression response (16), resulting in low levels of transcription of genes involved in respiration and in the tricarboxylic acid (TCA) cycle. In contrast, the Crabtree-negative P. stipitis exhibits predominantly respirative metabolism even at high glucose concentrations (33). These obvious differences in metabolic regulation between the two yeasts provide motivation for investigation of potential differences in the central carbon pathways.…”

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confidence: 99%

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Metabolic-Flux Profiling of the Yeasts Saccharomyces cerevisiae and Pichia stipitis

et al. 2003

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The so far largely uncharacterized central carbon metabolism of the yeast Pichia stipitis was explored in batch and glucose-limited chemostat cultures using metabolic-flux ratio analysis by nuclear magnetic resonance. The concomitantly characterized network of active metabolic pathways was compared to those identified in Saccharomyces cerevisiae, which led to the following conclusions. (i) There is a remarkably low use of the non-oxidative pentose phosphate (PP) pathway for glucose catabolism in S. cerevisiae when compared to P. stipitis batch cultures. (ii) Metabolism of P. stipitis batch cultures is fully respirative, which contrasts with the predominantly respiro-fermentative metabolic state of S. cerevisiae. (iii) Glucose catabolism in chemostat cultures of both yeasts is primarily oxidative. (iv) In both yeasts there is significant in vivo malic enzyme activity during growth on glucose. (v) The amino acid biosynthesis pathways are identical in both yeasts. The present investigation thus demonstrates the power of metabolic-flux ratio analysis for comparative profiling of central carbon metabolism in lower eukaryotes. Although not used for glucose catabolism in batch culture, we demonstrate that the PP pathway in S. cerevisiae has a generally high catabolic capacity by overexpressing the Escherichia coli transhydrogenase UdhA in phosphoglucose isomerase-deficient S. cerevisiae.The two yeasts Saccharomyces cerevisiae and Pichia stipitis exhibit fundamentally different modes of metabolic regulation in glucose-containing media. At high extracellular concentrations of glucose, one observes simultaneous fermentation and respiration (respiro-fermentative metabolism) in S. cerevisiae at high growth rates even under fully aerobic conditions (11,34,53). In Crabtree-positive yeasts, such as S. cerevisiae, elevated glucose concentrations induce the carbon catabolite repression response (16), resulting in low levels of transcription of genes involved in respiration and in the tricarboxylic acid (TCA) cycle. In contrast, the Crabtree-negative P. stipitis exhibits predominantly respirative metabolism even at high glucose concentrations (33). These obvious differences in metabolic regulation between the two yeasts provide motivation for investigation of potential differences in the central carbon pathways. Stable isotope labeling experiments appear to be a promising approach, since it reveals in vivo activity of pathways and reactions (10,43). Biosynthetically directed fractional (BDF) 13 C labeling of amino acids can provide comprehensive insight into central carbon metabolism, yielding a network of active pathways and quantification of intracellular flux ratios (36,37,44,46); its use in the present study promises to expand on the results of earlier labeling studies in yeast that focused on intermediates or products of individual pathways or reactions (14,17,20,22,41,57). BDF 13 C labeling is achieved by growing cells on mixtures of unlabeled and uniformly 13 C-labeled [U-13 C 6 ]glucose (40, 44). Using two-dimensional (...

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Section: Discussionsupporting

confidence: 74%

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confidence: 99%

Metabolic-Flux Profiling of the Yeasts Saccharomyces cerevisiae and Pichia stipitis

et al. 2003

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“…Under this growth condition, no ethanol production is detected (Passoth et al, 1996;Cho and Jeffries, 1999). mRNA was extracted using a Dynabeads mRNA Direct  kit (Dynal).…”

Section: Mrna Isolation and Rt-pcrmentioning

confidence: 96%

“…The list mainly encompasses yeasts that are unable to produce ethanol in the presence of fermentable sugars under strictly aerobic conditions (Crabtree-negative). Pichia stipitis has the characteristics of a Crabtree-negative yeast (Passoth et al, 1996;Jeffries and Shi, 1999) and it possesses both CYT and STO respiration systems in its mitochondria (Jeppsson et al, 1995;Shi et al, 1999;Shi, 2000). Oxygen limitation, rather than the increase of metabolites in the lower part of SHAM-sensitive alternative respiration in Pichia stipitis 1205 glycolysis, induces the onset of ethanol formation in P. stipitis (Bruinenberg et al, 1984;Alexander and Jeffries, 1990;Cho and Jeffries, 1999).…”

Section: Introductionmentioning

confidence: 99%

SHAM‐sensitive alternative respiration in the xylose‐metabolizing yeast Pichia stipitis

Shi

Cruz

Sherman

et al. 2002

Yeast

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SHAM-sensitive (STO) alternative respiration is present in the xylose-metabolizing, Crabtree-negative yeast, Pichia stipitis, but its pathway components and physiological roles during xylose metabolism are poorly understood. We cloned PsSTO1, which encodes the SHAM-sensitive terminal oxidase (PsSto1p), by genome walking from wild-type CBS 6054 and subsequently deleted PsSTO1 by targeted gene disruption. The resulting sto1-deletion mutant, FPL-Shi31, did not contain other isoforms of Sto protein that were detectable by Western blot analysis using an alternative oxidase monoclonal antibody raised against the Sto protein from Sauromatum guttatum. Levels of cytochromes b, c, c 1 and a·a 3 did not change in the sto1-mutant, which indicated that deleting PsSto1p did not alter the cytochrome pool. Interestingly, the sto1-deletion mutant stopped growing earlier than the parent and produced 20% more ethanol from xylose. Heterologous expression of PsSTO1 in Saccharomyces cerevisiae increased its total oxygen consumption rate and imparted cyanide-resistant oxygen uptake but did not enable growth on ethanol, indicating that PsSto1p is not coupled to ATP synthesis. We present evidence that the mitochondrial NADH dehydrogenase complex (Complex I) was present in wild-type CBS 6054 but was bypassed in the cells during xylose metabolism. Unexpectedly, deleting PsSto1p led to the use of Complex I in the mutant cells when xylose was the carbon source. We propose that the non-proton-translocating NAD(P)H dehydrogenases are linked to PsSto1p in xylose-metabolizing cells and that this non-ATP-generating route serves a regulatory function in the complex redox network of P. stipitis. Published in

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“…Xylose and glucose are not equal fermentations for many reasons, but increasing the capacity of P. stipitis for rapid xylose fermentation can greatly develop its usefulness in commercial applications. One of fermentative regulatory machinery is that S. cerevisiae regulates fermentation process by sensing the existence of glucose, but P. stipitis promotes fermentative activity in response to oxygen limitation [114][115][116][117]. Universal expression array analysis has shown special response patterns for rahamnose, cellobiose, arabinose, xylose and other lignocellulosic substrates.…”

Section: New Yeast For Lignocelluloses Bioconversionmentioning

confidence: 99%

Bioethanol a Microbial Biofuel Metabolite; New Insights of Yeasts Metabolic Engineering

et al. 2018

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Scarcity of the non-renewable energy sources, global warming, environmental pollution, and raising the cost of petroleum are the motive for the development of renewable, eco-friendly fuels production with low costs. Bioethanol production is one of the promising materials that can subrogate the petroleum oil, and it is considered recently as a clean liquid fuel or a neutral carbon. Diverse microorganisms such as yeasts and bacteria are able to produce bioethanol on a large scale, which can satisfy our daily needs with cheap and applicable methods. Saccharomyces cerevisiae and Pichia stipitis are two of the pioneer yeasts in ethanol production due to their abilities to produce a high amount of ethanol. The recent focus is directed towards lignocellulosic biomass that contains 30-50% cellulose and 20-40% hemicellulose, and can be transformed into glucose and fundamentally xylose after enzymatic hydrolysis. For this purpose, a number of various approaches have been used to engineer different pathways for improving the bioethanol production with simultaneous fermentation of pentose and hexoses sugars in the yeasts. These approaches include metabolic and flux analysis, modeling and expression analysis, followed by targeted deletions or the overexpression of key genes. In this review, we highlight and discuss the current status of yeasts genetic engineering for enhancing bioethanol production, and the conditions that influence bioethanol production.

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Peculiarities of the Regulation of Fermentation and Respiration in the Crabtree-Negative, Xylose-Fermenting Yeast Pichia stipitis

Cited by 36 publications

References 28 publications

Metabolic-Flux Profiling of the Yeasts Saccharomyces cerevisiae and Pichia stipitis

Metabolic-Flux Profiling of the Yeasts Saccharomyces cerevisiae and Pichia stipitis

SHAM‐sensitive alternative respiration in the xylose‐metabolizing yeast Pichia stipitis

Bioethanol a Microbial Biofuel Metabolite; New Insights of Yeasts Metabolic Engineering

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