TCA cycle-independent acetate metabolism via the glyoxylate cycle in Saccharomyces cerevisiae

Yeast ◽  
2010 ◽  
Vol 28 (2) ◽  
pp. 153-166 ◽  
Author(s):  
Yong Joo Lee ◽  
Jin Won Jang ◽  
Kyung Jin Kim ◽  
Pil Jae Maeng
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Glyoxylate Cycle ◽  
Tca Cycle ◽  
Acetate Metabolism ◽  
The Glyoxylate Cycle

scholarly journals Mutants of Saccharomyces cerevisiae with Defects in Acetate Metabolism: Isolation and Characterization of Acn− Mutants

Genetics ◽  
1996 ◽  
Vol 144 (1) ◽  
pp. 57-69 ◽  
Author(s):  
Mark T McCammon
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Glyoxylate Cycle ◽  
Tca Cycle ◽  
Acetate Metabolism ◽  
Yeast Saccharomyces Cerevisiae ◽  
Isolation And Characterization ◽  
Complex Process ◽  
Metabolic Interactions ◽  
Cellular Compartments ◽  
The Glyoxylate Cycle

Abstract The two carbon compounds, ethanol and acetate, can be oxidatively metabolized as well as assimilated into carbohydrate in the yeast Saccharomyces cerevisiae. The distribution of acetate metabolic enzymes among several cellular compartments, mitochondria, peroxisomes, and cytoplasm makes it an intriguing system to study complex metabolic interactions. To investigate the complex process of carbon catabolism and assimilation, mutants unable to grow on acetate were isolated. One hundred five Acn− (“Acetate Nonutilizing”) mutants were sorted into 21 complementation groups with an additional 20 single mutants. Five of the groups have defects in TCA cycle enzymes: MDH1, CIT1, ACO1, IDH1, and IDH2. A defect in RTG2, involved in the retrograde communication between the mitochondrion and the nucleus, was also identified. Four genes encode enzymes of the glyoxylate cycle and gluconeogenesis: ICL1, MLS1, MBH2, and PCK1. Five other genes appear to be defective in regulating metabolic activity since elevated levels of enzymes in several metabolic pathways, including the glyoxylate cycle, gluconeogenesis, and acetyl-coA metabolism, were detected in these mutants: ACN8, ACN9, ACNl7, ACN18, and ACN42. In summary, this analysis has identified at least 22 and as many as 41 different genes involved in acetate metabolism.

Acetate metabolism in Escherichia coli

1978 ◽  
Vol 24 (2) ◽  
pp. 149-153 ◽  
Author(s):  
T. M. Lakshmi ◽  
Robert B. Helling
Keyword(s):  
Isocitrate Dehydrogenase ◽  
Citrate Synthase ◽  
Isocitrate Lyase ◽  
Glyoxylate Cycle ◽  
Acetate Metabolism ◽  
Wild Type ◽  
Intermediary Metabolites ◽  
Lyase Activity ◽  
Acetate Medium ◽  
The Glyoxylate Cycle

Levels of several intermediary metabolites were measured in cells grown in acetate medium in order to test the hypothesis that the glyoxylate cycle is repressed by phosphoenolpyruvate (PEP). Wild-type cells had less PEP than either isocitrate dehydrogenase – deficient cells (which had greater isocitrate lyase activity than the wild type) or isocitrate dehydrogenase – deficient, citrate synthase – deficient cells (which are poorly inducible). Thus induction of the glyoxylate cycle is more complicated than a simple function of PEP concentration. No correlation between enzyme activity and the level of oxaloacetate, pyruvate, or citrate was found either. Citrate was synthesized in citrate synthase – deficient mutants, possibly via citrate lyase.

scholarly journals Engineering and Evolution of Methanol Assimilation in Saccharomyces cerevisiae

2019 ◽  
Author(s):  
Monica I. Espinosa ◽  
Ricardo A. Gonzalez-Garcia ◽  
Kaspar Valgepea ◽  
Manuel Plan ◽  
Colin Scott ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Yeast Extract ◽  
Glyoxylate Cycle ◽  
Carbon Assimilation ◽  
Tca Cycle ◽  
Ethanol Metabolism ◽  
Attractive Alternative ◽  
Chemical Production ◽  
Adaptive Laboratory Evolution ◽  
Methanol Yeast

AbstractMicrobial fermentation for chemical production is becoming more broadly adopted as an alternative to petrochemical refining. Fermentation typically relies on sugar as a feedstock, however, one-carbon compounds like methanol are an attractive alternative as they can be derived from organic waste and natural gas. This study focused on engineering methanol assimilation in the yeast Saccharomyces cerevisiae. Three methanol assimilation pathways were engineered and tested: a synthetic xylulose monophosphate (XuMP), a ‘hybrid’ methanol dehydrogenase-XuMP, and a bacterial ribulose monophosphate (RuMP) pathway, with the latter identified as the most effective at assimilating methanol. Additionally, 13C-methanol tracer analysis uncovered a native capacity for methanol assimilation in S. cerevisiae, which was optimized using Adaptive Laboratory Evolution. Three independent lineages selected in liquid methanol-yeast extract medium evolved premature stop codons in YGR067C, which encodes an uncharacterised protein that has a predicted DNA-binding domain with homology to the ADR1 transcriptional regulator. Adr1p regulates genes involved in ethanol metabolism and peroxisomal proliferation, suggesting YGR067C has a related function. When one of the evolved YGR067C mutations was reverse engineered into the parental CEN.PK113-5D strain, there were up to 5-fold increases in 13C-labelling of intracellular metabolites from 13C-labelled methanol when 0.1 % yeast extract was a co-substrate, and a 44 % increase in final biomass. Transcriptomics and proteomics revealed that the reconstructed YGR067C mutation results in down-regulation of genes in the TCA cycle, glyoxylate cycle, and gluconeogenesis, which would normally be up-regulated during growth on a non-fermentable carbon source. Combining the synthetic RuMP and XuMP pathways with the reconstructed Ygr067cp truncation led to further improvements in growth. These results identify a latent methylotrophic metabolism in S. cerevisiae and pave the way for further development of native and synthetic one-carbon assimilation pathways in this model eukaryote.

scholarly journals Anaplerotic reactions active during growth of Saccharomyces cerevisiae on glycerol

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Joeline Xiberras ◽  
Mathias Klein ◽  
Celina Prosch ◽  
Zahabiya Malubhoy ◽  
Elke Nevoigt
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Growth Rate ◽  
Specific Growth Rate ◽  
Pyruvate Carboxylase ◽  
Reference Strain ◽  
Specific Growth ◽  
Glyoxylate Cycle ◽  
Maximum Specific Growth Rate ◽  
The Glyoxylate Cycle ◽  
Anaplerotic Reactions

ABSTRACT Anaplerotic reactions replenish TCA cycle intermediates during growth. In Saccharomyces cerevisiae, pyruvate carboxylase and the glyoxylate cycle have been experimentally identified to be the main anaplerotic routes during growth on glucose (C6) and ethanol (C2), respectively. The current study investigates the importance of the two isoenzymes of pyruvate carboxylase (PYC1 and PYC2) and one of the key enzymes of the glyoxylate cycle (ICL1) for growth on glycerol (C3) as a sole carbon source. As the wild-type strains of the CEN.PK family are unable to grow in pure synthetic glycerol medium, a reverse engineered derivative showing a maximum specific growth rate of 0.14 h−1 was used as the reference strain. While the deletion of PYC1 reduced the maximum specific growth rate by about 38%, the deletion of PYC2 had no significant impact, neither in the reference strain nor in the pyc1Δ mutant. The deletion of ICL1 only marginally reduced growth of the reference strain but further decreased the growth rate of the pyc1 deletion strain by 20%. Interestingly, the triple deletion (pyc1Δ pyc2Δ icl1Δ) did not show any growth. Therefore, both the pyruvate carboxylase and the glyoxylate cycle are involved in anaplerosis during growth on glycerol.

scholarly journals Metabolic and Developmental Effects Resulting from Deletion of the citA Gene Encoding Citrate Synthase in Aspergillus nidulans

2010 ◽  
Vol 9 (4) ◽  
pp. 656-666 ◽  
Author(s):  
Sandra L. Murray ◽  
Michael J. Hynes
Keyword(s):  
Aspergillus Nidulans ◽  
Fluorescent Protein ◽  
Citrate Synthase ◽  
Glyoxylate Cycle ◽  
Single Gene ◽  
Carbon Sources ◽  
Tca Cycle ◽  
Poor Growth ◽  
Content Type ◽  
The Glyoxylate Cycle

ABSTRACT Citrate synthase is a central activity in carbon metabolism. It is required for the tricarboxylic acid (TCA) cycle, respiration, and the glyoxylate cycle. In Saccharomyces cerevisiae and Arabidopsis thaliana, there are mitochondrial and peroxisomal isoforms encoded by separate genes, while in Aspergillus nidulans, a single gene, citA, encodes a protein with predicted mitochondrial and peroxisomal targeting sequences (PTS). Deletion of citA results in poor growth on glucose but not on derepressing carbon sources, including those requiring the glyoxylate cycle. Growth on glucose is restored by a mutation in the creA carbon catabolite repressor gene. Methylcitrate synthase, required for propionyl-coenzyme A (CoA) metabolism, has previously been shown to have citrate synthase activity. We have been unable to construct the mcsAΔ citAΔ double mutant, and the expression of mcsA is subject to CreA-mediated carbon repression. Therefore, McsA can substitute for the loss of CitA activity. Deletion of citA does not affect conidiation or sexual development but results in delayed conidial germination as well as a complete loss of ascospores in fruiting bodies, which can be attributed to loss of meiosis. These defects are suppressed by the creA204 mutation, indicating that McsA activity can substitute for the loss of CitA. A mutation of the putative PTS1-encoding sequence in citA had no effect on carbon source utilization or development but did result in slower colony extension arising from single conidia or ascospores. CitA-green fluorescent protein (GFP) studies showed mitochondrial localization in conidia, ascospores, and hyphae. Peroxisomal localization was not detected. However, a very low and variable detection of punctate GFP fluorescence was sometimes observed in conidia germinated for 5 h when the mitochondrial targeting sequence was deleted.

scholarly journals Structure of glyoxysomal malate dehydrogenase (MDH3) from Saccharomyces cerevisiae

2018 ◽  
Vol 74 (10) ◽  
pp. 617-624 ◽  
Author(s):  
Shu Moriyama ◽  
Kazuya Nishio ◽  
Tsunehiro Mizushima
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Malate Dehydrogenase ◽  
Ternary Complex ◽  
Tricarboxylic Acid Cycle ◽  
Glyoxylate Cycle ◽  
Tricarboxylic Acid ◽  
Mitochondrial Enzyme ◽  
Open Conformation ◽  
Metabolism Enzyme ◽  
The Glyoxylate Cycle

Malate dehydrogenase (MDH), a carbohydrate and energy metabolism enzyme in eukaryotes, catalyzes the interconversion of malate to oxaloacetate (OAA) in conjunction with that of nicotinamide adenine dinucleotide (NAD+) to NADH. Three isozymes of MDH have been reported in Saccharomyces cerevisiae: MDH1, MDH2 and MDH3. MDH1 is a mitochondrial enzyme and a member of the tricarboxylic acid cycle, whereas MDH2 is a cytosolic enzyme that functions in the glyoxylate cycle. MDH3 is a glyoxysomal enzyme that is involved in the reoxidation of NADH, which is produced during fatty-acid β-oxidation. The affinity of MDH3 for OAA is lower than those of MDH1 and MDH2. Here, the crystal structures of yeast apo MDH3, the MDH3–NAD+ complex and the MDH3–NAD+–OAA ternary complex were determined. The structure of the ternary complex suggests that the active-site loop is in the open conformation, differing from the closed conformations in mitochondrial and cytosolic malate dehydrogenases.

scholarly journals Multiple Contributions of Peroxisomal Metabolic Function to Fungal Pathogenicity in Colletotrichum lagenarium

2006 ◽  
Vol 72 (9) ◽  
pp. 6345-6354 ◽  
Author(s):  
Makoto Asakura ◽  
Tetsuro Okuno ◽  
Yoshitaka Takano
Keyword(s):  
Host Plant ◽  
Isocitrate Lyase ◽  
Glyoxylate Cycle ◽  
Acetate Metabolism ◽  
Colletotrichum Lagenarium ◽  
Melanin Biosynthesis ◽  
Fungal Pathogenicity ◽  
Peroxisomal Targeting ◽  
The Glyoxylate Cycle ◽  
Host Invasion

ABSTRACT In Colletotrichum lagenarium, which is the causal agent of cucumber anthracnose, PEX6 is required for peroxisome biogenesis and appressorium-mediated infection. To verify the roles of peroxisome-associated metabolism in fungal pathogenicity, we isolated and functionally characterized ICL1 of C. lagenarium, which encodes isocitrate lyase involved in the glyoxylate cycle in peroxisomes. The icl1 mutants failed to utilize fatty acids and acetate for growth. Although Icl1 has no typical peroxisomal targeting signals, expression analysis of the GFP-Icl1 fusion protein indicated that Icl1 localizes in peroxisomes. These results indicate that the glyoxylate cycle that occurs inside the peroxisome is required for fatty acid and acetate metabolism for growth. Importantly, in contrast with the pex6 mutants that form nonmelanized appressoria, the icl1 mutants formed appressoria that were highly pigmented with melanin, suggesting that the glyoxylate cycle is not essential for melanin biosynthesis in appressoria. However, the icl1 mutants exhibited a severe reduction in virulence. Appressoria of the icl1 mutants failed to develop penetration hyphae in the host plant, suggesting that ICL1 is involved in host invasion. The addition of glucose partially restored virulence of the icl1 mutant. Heat shock treatment of the host plant also enabled the icl1 mutants to develop lesions, implying that the infection defect of the icl1 mutant is associated with plant defense. Together with the requirement of PEX6 for appressorial melanization, our findings suggest that peroxisomal metabolic pathways play functional roles in appressorial melanization and subsequent host invasion steps, and the latter step requires the glyoxylate cycle.

scholarly journals A Transcriptional Switch in the Expression of Yeast Tricarboxylic Acid Cycle Genes in Response to a Reduction or Loss of Respiratory Function

1999 ◽  
Vol 19 (10) ◽  
pp. 6720-6728 ◽  
Author(s):  
Zhengchang Liu ◽  
Ronald A. Butow
Keyword(s):  
Binding Site ◽  
Respiratory Function ◽  
Leucine Zipper ◽  
Tricarboxylic Acid Cycle ◽  
Glyoxylate Cycle ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Helix Loop Helix ◽  
Expression Of Genes ◽  
The Glyoxylate Cycle

ABSTRACT The Hap2,3,4,5p transcription complex is required for expression of many mitochondrial proteins that function in electron transport and the tricarboxylic acid (TCA) cycle. We show that as the cells’ respiratory function is reduced or eliminated, the expression of four TCA cycle genes, CIT1, ACO1, IDH1, andIDH2, switches from HAP control to control by three genes, RTG1, RTG2, and RTG3. The expression of four additional TCA cycle genes downstream ofIDH1 and IDH2 is independent of theRTG genes. We have previously shown that theRTG genes control the retrograde pathway, defined as a change in the expression of a subset of nuclear genes, e.g., the glyoxylate cycle CIT2 gene, in response to changes in the functional state of mitochondria. We show that thecis-acting sequence controlling RTG-dependent expression of CIT1 includes an R box element, GTCAC, located 70 bp upstream of the Hap2,3,4,5p binding site in theCIT1 upstream activation sequence. The R box is a binding site for Rtg1p-Rtg3p, a heterodimeric, basic helix-loop-helix/leucine zipper transcription factor complex. We propose that in cells with compromised mitochondrial function, the RTG genes take control of the expression of genes leading to the synthesis of α-ketoglutarate to ensure that sufficient glutamate is available for biosynthetic processes and that increased flux of the glyoxylate cycle, via elevated CIT2 expression, provides a supply of metabolites entering the TCA cycle sufficient to support anabolic pathways. Glutamate is a potent repressor of RTG-dependent expression of genes encoding both mitochondrial and nonmitochondrial proteins, suggesting that it is a specific feedback regulator of the RTG system.

scholarly journals Studies on the Regulation and Localization of the Glyoxylate Cycle Enzymes in Saccharomyces cerevisiae

2005 ◽  
Vol 10 (1) ◽  
pp. 83-89 ◽  
Author(s):  
W. Dduntze ◽  
D. Neumann ◽  
W. Atzpodien ◽  
H. Holzer ◽  
J. M. Gancedo
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Glyoxylate Cycle ◽  
The Glyoxylate Cycle
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