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Cell Oct 2018Acetate is a major nutrient that supports acetyl-coenzyme A (Ac-CoA) metabolism and thus lipogenesis and protein acetylation. However, its source is unclear. Here, we...
Acetate is a major nutrient that supports acetyl-coenzyme A (Ac-CoA) metabolism and thus lipogenesis and protein acetylation. However, its source is unclear. Here, we report that pyruvate, the end product of glycolysis and key node in central carbon metabolism, quantitatively generates acetate in mammals. This phenomenon becomes more pronounced in the context of nutritional excess, such as during hyperactive glucose metabolism. Conversion of pyruvate to acetate occurs through two mechanisms: (1) coupling to reactive oxygen species (ROS) and (2) neomorphic enzyme activity from keto acid dehydrogenases that enable function as pyruvate decarboxylases. Further, we demonstrate that de novo acetate production sustains Ac-CoA pools and cell proliferation in limited metabolic environments, such as during mitochondrial dysfunction or ATP citrate lyase (ACLY) deficiency. By virtue of de novo acetate production being coupled to mitochondrial metabolism, there are numerous possible regulatory mechanisms and links to pathophysiology.
Topics: ATP Citrate (pro-S)-Lyase; Acetates; Acetyl Coenzyme A; Acetylation; Animals; Female; Glucose; Glycolysis; Lipogenesis; Male; Mammals; Mice; Mice, Inbred C57BL; Mitochondria; Oxidoreductases; Pyruvate Decarboxylase; Pyruvic Acid; Reactive Oxygen Species
PubMed: 30245009
DOI: 10.1016/j.cell.2018.08.040 -
BMC Research Notes May 2021Zymomonas mobilis is an alpha-proteobacterium with a rapid ethanologenic pathway, involving Entner-Doudoroff (E-D) glycolysis, pyruvate decarboxylase (Pdc) and two...
OBJECTIVE
Zymomonas mobilis is an alpha-proteobacterium with a rapid ethanologenic pathway, involving Entner-Doudoroff (E-D) glycolysis, pyruvate decarboxylase (Pdc) and two alcohol dehydrogenase (ADH) isoenzymes. Pyruvate is the end-product of the E-D pathway and the substrate for Pdc. Construction and study of Pdc-deficient strains is of key importance for Z. mobilis metabolic engineering, because the pyruvate node represents the central branching point, most novel pathways divert from ethanol synthesis. In the present work, we examined the aerobic metabolism of a strain with partly inactivated Pdc.
RESULTS
Relative to its parent strain the mutant produced more pyruvate. Yet, it also yielded more acetaldehyde, the product of the Pdc reaction and the substrate for ADH, although the bulk ADH activity was similar in both strains, while the Pdc activity in the mutant was reduced by half. Simulations with the kinetic model of Z. mobilis E-D pathway indicated that, for the observed acetaldehyde to ethanol production ratio in the mutant, the ratio between its respiratory NADH oxidase and ADH activities should be significantly higher, than the measured values. Implications of this finding for the directionality of the ADH isoenzyme operation in vivo and interactions between ADH and Pdc are discussed.
Topics: Alcohol Dehydrogenase; Metabolic Engineering; Pyruvate Decarboxylase; Respiration; Zymomonas
PubMed: 34049566
DOI: 10.1186/s13104-021-05625-5 -
PloS One 2023Understanding metabolism in the pathogen Candida glabrata is key to identifying new targets for antifungals. The thiamine biosynthetic (THI) pathway is partially...
Understanding metabolism in the pathogen Candida glabrata is key to identifying new targets for antifungals. The thiamine biosynthetic (THI) pathway is partially defective in C. glabrata, but the transcription factor CgPdc2 upregulates some thiamine biosynthetic and transport genes. One of these genes encodes a recently evolved thiamine pyrophosphatase (CgPMU3) that is critical for accessing external thiamine. Here, we demonstrate that CgPdc2 primarily regulates THI genes. In Saccharomyces cerevisiae, Pdc2 regulates both THI and pyruvate decarboxylase (PDC) genes, with PDC proteins being a major thiamine sink. Deletion of PDC2 is lethal in S. cerevisiae in standard growth conditions, but not in C. glabrata. We uncover cryptic cis elements in C. glabrata PDC promoters that still allow for regulation by ScPdc2, even when that regulation is not apparent in C. glabrata. C. glabrata lacks Thi2, and it is likely that inclusion of Thi2 into transcriptional regulation in S. cerevisiae allows for a more complex regulation pattern and regulation of THI and PDC genes. We present evidence that Pdc2 functions independent of Thi2 and Thi3 in both species. The C-terminal activation domain of Pdc2 is intrinsically disordered and critical for species differences. Truncation of the disordered domains leads to a gradual loss of activity. Through a series of cross species complementation assays of transcription, we suggest that there are multiple Pdc2-containing complexes, and C. glabrata appears to have the simplest requirement set for THI genes, except for CgPMU3. CgPMU3 has different cis requirements, but still requires Pdc2 and Thi3 to be upregulated by thiamine starvation. We identify the minimal region sufficient for thiamine regulation in CgTHI20, CgPMU3, and ScPDC5 promoters. Defining the cis and trans requirements for THI promoters should lead to an understanding of how to interrupt their upregulation and provide targets in metabolism for antifungals.
Topics: Saccharomyces cerevisiae; Candida glabrata; Transcription Factors; Fungal Proteins; Pyruvate Decarboxylase; Thiamine; Carboxy-Lyases; Promoter Regions, Genetic; Intrinsically Disordered Proteins; Gene Expression Regulation, Fungal
PubMed: 37285346
DOI: 10.1371/journal.pone.0286744 -
The FEBS Journal Dec 2013Thiamine diphosphate-dependent enzymes are broadly distributed in all organisms, and they catalyse a broad range of C-C bond forming and breaking reactions. Enzymes... (Review)
Review
Thiamine diphosphate-dependent enzymes are broadly distributed in all organisms, and they catalyse a broad range of C-C bond forming and breaking reactions. Enzymes belonging to the structural families of decarboxylases and transketolases have been particularly well investigated concerning their substrate range, mechanism of stereoselective carboligation and carbolyase reaction. Both structurally different enzyme families differ also in stereoselectivity: enzymes from the decarboxylase family are predominantly R-selective, whereas those from the transketolase family are S-selective. In recent years a key focus of our studies has been on stereoselective benzoin condensation-like 1,2-additions. Meanwhile, several S-selective variants of pyruvate decarboxylase, benzoylformate decarboxylase and 2-succinyl-5-enolpyruvyl-6-hydroxy-3-cyclohexene-1-carboxylate (SEPHCHC) synthase as well as R-selective transketolase variants were created that allow access to a broad range of enantiocomplementary α-hydroxyketones and α,α'-dihydroxyketones. This review covers recent studies and the mechanistic understanding of stereoselective C-C bond forming thiamine diphosphate-dependent enzymes, which has been guided by structure-function analyses based on mutagenesis studies and from influences of different substrates and organic co-solvents on stereoselectivity.
Topics: Animals; Carboxy-Lyases; Catalysis; Humans; Stereoisomerism; Thiamine Pyrophosphate; Transketolase
PubMed: 24034356
DOI: 10.1111/febs.12496 -
BMC Structural Biology Nov 2014Bacterial pyruvate decarboxylases (PDC) are rare. Their role in ethanol production and in bacterially mediated ethanologenic processes has, however, ensured a continued...
BACKGROUND
Bacterial pyruvate decarboxylases (PDC) are rare. Their role in ethanol production and in bacterially mediated ethanologenic processes has, however, ensured a continued and growing interest. PDCs from Zymomonas mobilis (ZmPDC), Zymobacter palmae (ZpPDC) and Sarcina ventriculi (SvPDC) have been characterized and ZmPDC has been produced successfully in a range of heterologous hosts. PDCs from the Acetobacteraceae and their role in metabolism have not been characterized to the same extent. Examples include Gluconobacter oxydans (GoPDC), G. diazotrophicus (GdPDC) and Acetobacter pasteutrianus (ApPDC). All of these organisms are of commercial importance.
RESULTS
This study reports the kinetic characterization and the crystal structure of a PDC from Gluconacetobacter diazotrophicus (GdPDC). Enzyme kinetic analysis indicates a high affinity for pyruvate (K M 0.06 mM at pH 5), high catalytic efficiencies (1.3 • 10(6) M(-1) • s(-1) at pH 5), pHopt of 5.5 and Topt at 45°C. The enzyme is not thermostable (T½ of 18 minutes at 60°C) and the calculated number of bonds between monomers and dimers do not give clear indications for the relatively lower thermostability compared to other PDCs. The structure is highly similar to those described for Z. mobilis (ZmPDC) and A. pasteurianus PDC (ApPDC) with a rmsd value of 0.57 Å for Cα when comparing GdPDC to that of ApPDC. Indole-3-pyruvate does not serve as a substrate for the enzyme. Structural differences occur in two loci, involving the regions Thr341 to Thr352 and Asn499 to Asp503.
CONCLUSIONS
This is the first study of the PDC from G. diazotrophicus (PAL5) and lays the groundwork for future research into its role in this endosymbiont. The crystal structure of GdPDC indicates the enzyme to be evolutionarily closely related to homologues from Z. mobilis and A. pasteurianus and suggests strong selective pressure to keep the enzyme characteristics in a narrow range. The pH optimum together with reduced thermostability likely reflect the host organisms niche and conditions under which these properties have been naturally selected for. The lack of activity on indole-3-pyruvate excludes this decarboxylase as the enzyme responsible for indole acetic acid production in G. diazotrophicus.
Topics: Amino Acids; Bacterial Proteins; Crystallography, X-Ray; Gluconacetobacter; Models, Molecular; Phylogeny; Protein Conformation; Protein Structure, Quaternary; Protein Structure, Tertiary; Pyruvate Decarboxylase; Sarcina; Sequence Homology, Amino Acid; Substrate Specificity; Zymomonas
PubMed: 25369873
DOI: 10.1186/s12900-014-0021-1 -
Journal of Applied Microbiology Jul 2015Ethanol production directly from CO2 , utilizing genetically engineered photosynthetic cyanobacteria as a biocatalyst, offers significant potential as a renewable and... (Review)
Review
Ethanol production directly from CO2 , utilizing genetically engineered photosynthetic cyanobacteria as a biocatalyst, offers significant potential as a renewable and sustainable source of biofuel. Despite the current absence of a commercially successful production system, significant resources have been deployed to realize this goal. Utilizing the pyruvate decarboxylase from Zymomonas species, metabolically derived pyruvate can be converted to ethanol. This review of both peer-reviewed and patent literature focuses on the genetic modifications utilized for metabolic engineering and the resultant effect on ethanol yield. Gene dosage, induced expression and cassette optimizat-ion have been analyzed to optimize production, with production rates of 0·1-0·5 g L(-1) day(-1) being achieved. The current 'toolbox' of molecular manipulations and future directions focusing on applicability, addressing the primary challenges facing commercialization of cyanobacterial technologies are discussed.
Topics: Autotrophic Processes; Biofuels; Cyanobacteria; Ethanol; Industrial Microbiology; Photosynthesis
PubMed: 25865951
DOI: 10.1111/jam.12821 -
Current Opinion in Chemical Biology Dec 2020Saccharomyces cerevisiae, Baker's yeast, is the industrial workhorse for producing ethanol and the subject of substantial metabolic engineering research in both industry... (Review)
Review
Saccharomyces cerevisiae, Baker's yeast, is the industrial workhorse for producing ethanol and the subject of substantial metabolic engineering research in both industry and academia. S. cerevisiae has been used to demonstrate production of a wide range of chemical products from glucose. However, in many cases, the demonstrations report titers and yields that fall below thresholds for industrial feasibility. Ethanol synthesis is a central part of S. cerevisiae metabolism, and redirecting flux to other products remains a barrier to industrialize strains for producing other molecules. Removing ethanol producing pathways leads to poor fitness, such as impaired growth on glucose. Here, we review metabolic engineering efforts aimed at restoring growth in non-ethanol producing strains with emphasis on relieving glucose repression associated with the Crabtree effect and rewiring metabolism to provide access to critical cellular building blocks. Substantial progress has been made in the past decade, but many opportunities for improvement remain.
Topics: Acetyl Coenzyme A; Ethanol; Glucose; Industrial Microbiology; Metabolic Engineering; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 33032255
DOI: 10.1016/j.cbpa.2020.08.005 -
Computational and Structural... 2023Short-chain fatty acids (SCFAs) exhibit anticancer activity in cellular and animal models of colon cancer. Acetate, propionate, and butyrate are the three major SCFAs...
Short-chain fatty acids (SCFAs) exhibit anticancer activity in cellular and animal models of colon cancer. Acetate, propionate, and butyrate are the three major SCFAs produced from dietary fiber by gut microbiota fermentation and have beneficial effects on human health. Most previous studies on the antitumor mechanisms of SCFAs have focused on specific metabolites or genes involved in antitumor pathways, such as reactive oxygen species (ROS) biosynthesis. In this study, we performed a systematic and unbiased analysis of the effects of acetate, propionate, and butyrate on ROS levels and metabolic and transcriptomic signatures at physiological concentrations in human colorectal adenocarcinoma cells. We observed significantly elevated levels of ROS in the treated cells. Furthermore, significantly regulated signatures were involved in overlapping pathways at metabolic and transcriptomic levels, including ROS response and metabolism, fatty acid transport and metabolism, glucose response and metabolism, mitochondrial transport and respiratory chain complex, one-carbon metabolism, amino acid transport and metabolism, and glutaminolysis, which are directly or indirectly linked to ROS production. Additionally, metabolic and transcriptomic regulation occurred in a SCFAs types-dependent manner, with an increasing degree from acetate to propionate and then to butyrate. This study provides a comprehensive analysis of how SCFAs induce ROS production and modulate metabolic and transcriptomic levels in colon cancer cells, which is vital for understanding the mechanisms of the effects of SCFAs on antitumor activity in colon cancer.
PubMed: 36874158
DOI: 10.1016/j.csbj.2023.02.022 -
Plant Signaling & Behavior 2014This review focuses on the energy metabolism during pollen maturation and tube growth and updates current knowledge. Pollen tube growth is essential for male... (Review)
Review
This review focuses on the energy metabolism during pollen maturation and tube growth and updates current knowledge. Pollen tube growth is essential for male reproductive success and extremely fast. Therefore, pollen development and tube growth are high energy-demanding processes. During the last years, various publications (including research papers and reviews) emphasize the importance of mitochondrial respiration and fermentation during male gametogenesis and pollen tube elongation. These pathways obviously contribute to satisfy the high energy demand, and there are many studies which suggest that respiration and fermentation are the only pathways to generate the needed energy. Here, we review data which show for the first time that in addition plastidial glycolysis and the balancing of the ATP/NAD(P)H ratio (by malate valves and NAD(+) biosynthesis) contribute to satisfy the energy demand during pollen development. Although the importance of energy generation by plastids was discounted during the last years (possibly due to the controversial opinion about their existence in pollen grains and pollen tubes), the available data underline their prime role during pollen maturation and tube growth.
Topics: Energy Metabolism; Fermentation; Mitochondria; Oxidation-Reduction; Plastids; Pollen Tube
PubMed: 25482752
DOI: 10.4161/15592324.2014.977200 -
Plants (Basel, Switzerland) Aug 2023Kiwifruit ( spp.) is susceptible to waterlogging stress. Although abundant wild germplasm resources exist among plants for improving the waterlogging tolerance of...
Kiwifruit ( spp.) is susceptible to waterlogging stress. Although abundant wild germplasm resources exist among plants for improving the waterlogging tolerance of kiwifruit cultivars, the underlying mechanisms remain largely unknown. Here, a comparative study was undertaken using one wild germplasm, Maorenshen ( Dunn, MRS), and one cultivar, Miliang-1 ( var. (A.Chev.) A.Chev. cv. Miliang-1, ML). Under stress, the ML plantlets were seriously damaged with wilted chlorotic leaves and blackened rotten roots, whereas the symptoms of injury in the MRS plantlets were much fewer, along with higher photosynthetic rates, chlorophyll fluorescence characteristics and root activity under stress conditions. However, neither aerenchyma in the root nor adventitious roots appeared in both germplasms upon stress exposure. The activities of pyruvate decarboxylase (PDC) and alcohol dehydrogenase (ADH), as well as their transcript levels, were constitutively higher in MRS than those in ML under both normal and stress conditions. Waterlogging stress significantly enhanced the PDC and ADH enzyme activities in both germplasms, which were 60.8% and 22.4% higher in the MRS roots than those in the ML roots under waterlogging stress, respectively. Moreover, MRS displayed higher activities of antioxidant enzymes, including SOD, CAT, and APX, as well as DPPH-radical scavenging ability, and decreased HO and MDA accumulation under both normal and stress conditions. Our findings suggest that the waterlogging tolerance of the wild germplasm was associated with high PDC and ADH, as well as antioxidant ability.
PubMed: 37571025
DOI: 10.3390/plants12152872