Insights into the unique carboxylation reactions in the metabolism of propylene and acetone

2020 ◽  
Vol 477 (11) ◽  
pp. 2027-2038
Author(s):  
Florence Mus ◽  
Hsin-Hua Wu ◽  
Alexander B. Alleman ◽  
Krista A. Shisler ◽  
Oleg A. Zadvornyy ◽  
...  
Keyword(s):  
Tricarboxylic Acid Cycle ◽  
Single Step ◽  
Natural Environments ◽  
Central Metabolism ◽  
Acetyl Coenzyme A ◽  
Carboxylating Enzymes ◽  
Intermediate Metabolites ◽  
Disulfide Oxidoreductase ◽  
Anthropogenic Processes ◽  
Hydrolysis Of

Alkenes and ketones are two classes of ubiquitous, toxic organic compounds in natural environments produced in several biological and anthropogenic processes. In spite of their toxicity, these compounds are utilized as primary carbon and energy sources or are generated as intermediate metabolites in the metabolism of other compounds by many diverse bacteria. The aerobic metabolism of some of the smallest and most volatile of these compounds (propylene, acetone, isopropanol) involves novel carboxylation reactions resulting in a common product acetoacetate. Propylene is metabolized in a four-step pathway involving five enzymes where the penultimate step is a carboxylation reaction catalyzed by a unique disulfide oxidoreductase that couples reductive cleavage of a thioether linkage with carboxylation to produce acetoacetate. The carboxylation of isopropanol begins with conversion to acetone via an alcohol dehydrogenase. Acetone is converted to acetoacetate in a single step by an acetone carboxylase which couples the hydrolysis of MgATP to the activation of both acetone and bicarbonate, generating highly reactive intermediates that are condensed into acetoacetate at a Mn2+ containing the active site. Acetoacetate is then utilized in central metabolism where it is readily converted to acetyl-coenzyme A and subsequently converted into biomass or utilized in energy metabolism via the tricarboxylic acid cycle. This review summarizes recent structural and biochemical findings that have contributed significant insights into the mechanism of these two unique carboxylating enzymes.

scholarly journals Proteogenomic Monitoring of Geobacter Physiology during Stimulated Uranium Bioremediation

2009 ◽  
Vol 75 (20) ◽  
pp. 6591-6599 ◽  
Author(s):  
Michael J. Wilkins ◽  
Nathan C. VerBerkmoes ◽  
Kenneth H. Williams ◽  
Stephen J. Callister ◽  
Paula J. Mouser ◽  
...  
Keyword(s):  
Tricarboxylic Acid Cycle ◽  
Protein Abundance ◽  
Central Metabolism ◽  
Community Members ◽  
Microbial Physiology ◽  
Acetyl Coenzyme A ◽  
Chromatography Tandem Mass Spectrometry ◽  
The Status ◽  
Liquid Chromatography Tandem Mass

ABSTRACT Implementation of uranium bioremediation requires methods for monitoring the membership and activities of the subsurface microbial communities that are responsible for reduction of soluble U(VI) to insoluble U(IV). Here, we report a proteomics-based approach for simultaneously documenting the strain membership and microbial physiology of the dominant Geobacter community members during in situ acetate amendment of the U-contaminated Rifle, CO, aquifer. Three planktonic Geobacter-dominated samples were obtained from two wells down-gradient of acetate addition. Over 2,500 proteins from each of these samples were identified by matching liquid chromatography-tandem mass spectrometry spectra to peptides predicted from seven isolate Geobacter genomes. Genome-specific peptides indicate early proliferation of multiple M21 and Geobacter bemidjiensis-like strains and later possible emergence of M21 and G. bemidjiensis-like strains more closely related to Geobacter lovleyi. Throughout biostimulation, the proteome is dominated by enzymes that convert acetate to acetyl-coenzyme A and pyruvate for central metabolism, while abundant peptides matching tricarboxylic acid cycle proteins and ATP synthase subunits were also detected, indicating the importance of energy generation during the period of rapid growth following the start of biostimulation. Evolving Geobacter strain composition may be linked to changes in protein abundance over the course of biostimulation and may reflect changes in metabolic functioning. Thus, metagenomics-independent community proteogenomics can be used to diagnose the status of the subsurface consortia upon which remediation biotechnology relies.

scholarly journals Oxidative Stress-Induced Alteration of Plant Central Metabolism

Life ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 304
Author(s):  
Tatyana Savchenko ◽  
Konstantin Tikhonov
Keyword(s):  
Oxidative Stress ◽  
Stress Responses ◽  
Tricarboxylic Acid Cycle ◽  
Strong Dependence ◽  
Plant Stress ◽  
Metabolite Profile ◽  
Agricultural Practice ◽  
Central Metabolism ◽  
Metabolic Markers ◽  
Sugar Derivatives

Oxidative stress is an integral component of various stress conditions in plants, and this fact largely determines the substantial overlap in physiological and molecular responses to biotic and abiotic environmental challenges. In this review, we discuss the alterations in central metabolism occurring in plants experiencing oxidative stress. To focus on the changes in metabolite profile associated with oxidative stress per se, we primarily analyzed the information generated in the studies based on the exogenous application of agents, inducing oxidative stress, and the analysis of mutants displaying altered oxidative stress response. Despite of the significant variation in oxidative stress responses among different plant species and tissues, the dynamic and transient character of stress-induced changes in metabolites, and the strong dependence of metabolic responses on the intensity of stress, specific characteristic changes in sugars, sugar derivatives, tricarboxylic acid cycle metabolites, and amino acids, associated with adaptation to oxidative stress have been detected. The presented analysis of the available data demonstrates the oxidative stress-induced redistribution of metabolic fluxes targeted at the enhancement of plant stress tolerance through the prevention of ROS accumulation, maintenance of the biosynthesis of indispensable metabolites, and production of protective compounds. This analysis provides a theoretical basis for the selection/generation of plants with improved tolerance to oxidative stress and the development of metabolic markers applicable in research and routine agricultural practice.

Determination of gluconeogenesis in vivo with 14C-labeled substrates

1985 ◽  
Vol 248 (4) ◽  
pp. R391-R399 ◽  
Author(s):  
J. Katz
Keyword(s):  
Pyruvate Carboxylase ◽  
Tricarboxylic Acid Cycle ◽  
Specific Radioactivity ◽  
Tricarboxylic Acid ◽  
Pyruvate Metabolism ◽  
Acetyl Coenzyme A ◽  
Glucose Carbon ◽  
Labeling Pattern

A mitochondrial model of gluconeogenesis and the tricarboxylic acid cycle, where pyruvate is metabolized via pyruvate carboxylase and pyruvate dehydrogenase, and pyruvate kinase is examined. The effect of the rate of tricarboxylic acid flux and the rates of the three reactions of pyruvate metabolism on the labeling patterns from [14C]pyruvate and [24C]acetate are analyzed. Expressions describing the specific radioactivities and 14C distribution in glucose as a function of these rates are derived. Specific radioactivities and isotopic patterns depend markedly on the ratio of the rates of pyruvate carboxylation and decarboxylation to the rate of citrate synthesis, but the effect of phosphoenolpyruvate hydrolysis is minor. The effects of these rates on 1) specific radioactivity of phosphoenolpyruvate, 2) labeling pattern in glucose, and 3) contribution of pyruvate, acetyl-coenzyme A, and CO2 to glucose carbon are illustrated. To determine the contribution of lactate or alanine to gluconeogenesis, experiments with two compounds labeled in different carbons are required. Methods in current use to correct for the dilution of 14C in gluconeogenesis from [14C]pyruvate are shown to be erroneous. The experimental design and techniques to determine gluconeogenesis from 14C-labeled precursors are presented and illustrated with numerical examples.

scholarly journals Bacterial cell cycle control by citrate synthase independent of enzymatic activity

2019 ◽  
Author(s):  
Matthieu Bergé ◽  
Julian Pezzatti ◽  
Víctor González-Ruiz ◽  
Laurence Degeorges ◽  
Serge Rudaz ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Citrate Synthase ◽  
Caulobacter Crescentus ◽  
Tricarboxylic Acid Cycle ◽  
Cell Cycle Control ◽  
Tca Cycle ◽  
Cell Cycle Regulators ◽  
Central Metabolism ◽  
Cycle Enzyme

ABSTRACTCoordination of cell cycle progression with central metabolism is fundamental to all cell types and likely underlies differentiation into dispersal cells in bacteria. How central metabolism is monitored to regulate cell cycle functions is poorly understood. A forward genetic selection for cell cycle regulators in the polarized alpha-proteobacterium Caulobacter crescentus unearthed the uncharacterized CitA citrate synthase, a TCA (tricarboxylic acid) cycle enzyme, as unprecedented checkpoint regulator of the G1→S transition. We show that loss of the CitA protein provokes a (p)ppGpp alarmone-dependent G1-phase arrest without apparent metabolic or energy insufficiency. While S-phase entry is still conferred when CitA is rendered catalytically inactive, the paralogous CitB citrate synthase has no overt role other than sustaining TCA cycle activity when CitA is absent. With eukaryotic citrate synthase paralogs known to fulfill regulatory functions, our work extends the moonlighting paradigm to citrate synthase coordinating central (TCA) metabolism with development and perhaps antibiotic tolerance in bacteria.

scholarly journals Substrate-dependent CO2 fixation in heterotrophic bacteria revealed by stable isotope labelling

2020 ◽  
Vol 96 (6) ◽  
Author(s):  
Marina Spona-Friedl ◽  
Alexander Braun ◽  
Claudia Huber ◽  
Wolfgang Eisenreich ◽  
Christian Griebler ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Heterotrophic Bacteria ◽  
Tricarboxylic Acid Cycle ◽  
Carbon Sources ◽  
Organic Substrate ◽  
Gas Chromatography Mass Spectrometry ◽  
Logarithmic Phase ◽  
Natural Environments ◽  
Organic Substrates ◽  
Stable Isotope Labelling

ABSTRACT Virtually all heterotrophs incorporate carbon dioxide by anaplerotic fixation. Little explored, however, is the interdependency of pathways and rates of CO2fixation on the concurrent usage of organic substrate(s). Potentially, this could reveal which substrates out of a pool of dissolved organic carbon are utilised by environmental microorganisms. To explore this possibility, Bacillus subtilis W23 was grown in a minimal medium with normalised amounts of either glucose, lactate or malate as only organic substrates, each together with 1 g/L NaH13CO3. Incorporation of H13CO3− was traced by elemental analysis-isotope ratio mass spectrometry of biomass and gas chromatography-mass spectrometry of protein-derived amino acids. Until the late logarithmic phase, 13C incorporation into the tricarboxylic acid cycle increased with time and occurred via [4–13C]oxaloacetate formed by carboxylation of pyruvate. The levels of 13C incorporation were highest for growth on glucose and lowest on malate. Incorporation of 13C into gluconeogenesis products was mainly detected in the lactate and malate experiment, whereas glucose down-regulated this path. A proof-of-principle study with a natural groundwater community confirmed the ability to determine incorporation from H13CO3− by natural communities leading to specific labelling patterns. This underlines the potential of the labelling approach to characterise carbon sources of heterotrophic microorganisms in their natural environments.

scholarly journals Conversion of 4-Hydroxybutyrate to Acetyl Coenzyme A and Its Anapleurosis in the Metallosphaera sedula 3-Hydroxypropionate/4-Hydroxybutyrate Carbon Fixation Pathway

2014 ◽  
Vol 80 (8) ◽  
pp. 2536-2545 ◽  
Author(s):  
Aaron B. Hawkins ◽  
Michael W. W. Adams ◽  
Robert M. Kelly
Keyword(s):  
Carbon Fixation ◽  
Coenzyme A ◽  
Flux Analysis ◽  
Central Metabolism ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Content Type ◽  
Metallosphaera Sedula ◽  
Acetyl Coenzyme ◽  
Coa Ligase

ABSTRACTThe extremely thermoacidophilic archaeonMetallosphaera sedula(optimum growth temperature, 73°C, pH 2.0) grows chemolithoautotrophically on metal sulfides or molecular hydrogen by employing the 3-hydroxypropionate/4-hydroxybutyrate (3HP/4HB) carbon fixation cycle. This cycle adds two CO2molecules to acetyl coenzyme A (acetyl-CoA) to generate 4HB, which is then rearranged and cleaved to form two acetyl-CoA molecules. Previous metabolic flux analysis showed that two-thirds of central carbon precursor molecules are derived from succinyl-CoA, which is oxidized to malate and oxaloacetate. The remaining one-third is apparently derived from acetyl-CoA. As such, the steps beyond succinyl-CoA are essential for completing the carbon fixation cycle and for anapleurosis of acetyl-CoA. Here, the final four enzymes of the 3HP/4HB cycle, 4-hydroxybutyrate-CoA ligase (AMP forming) (Msed_0406), 4-hydroxybutyryl-CoA dehydratase (Msed_1321), crotonyl-CoA hydratase/(S)-3-hydroxybutyryl-CoA dehydrogenase (Msed_0399), and acetoacetyl-CoA β-ketothiolase (Msed_0656), were produced recombinantly inEscherichia coli, combinedin vitro, and shown to convert 4HB to acetyl-CoA. Metabolic pathways connecting CO2fixation and central metabolism were examined using a gas-intensive bioreactor system in whichM. sedulawas grown under autotrophic (CO2-limited) and heterotrophic conditions. Transcriptomic analysis revealed the importance of the 3HP/4HB pathway in supplying acetyl-CoA to anabolic pathways generating intermediates inM. sedulametabolism. The results indicated that flux between the succinate and acetyl-CoA branches in the 3HP/4HB pathway is governed by 4-hydroxybutyrate-CoA ligase, possibly regulated posttranslationally by the protein acetyltransferase (Pat)/Sir2-dependent system. Taken together, this work confirms the final four steps of the 3HP/4HB pathway, thereby providing the framework for examining connections between CO2fixation and central metabolism inM. sedula.

scholarly journals Holistic bioengineering: rewiring central metabolism for enhanced bioproduction

2017 ◽  
Vol 474 (23) ◽  
pp. 3935-3950 ◽  
Author(s):  
Selçuk Aslan ◽  
Elad Noor ◽  
Arren Bar-Even
Keyword(s):  
Metabolic Network ◽  
Metabolic Pathways ◽  
Tricarboxylic Acid Cycle ◽  
Holistic Approach ◽  
Living Organism ◽  
Tricarboxylic Acid ◽  
High Availability ◽  
Biosynthesis Pathway ◽  
Central Metabolism ◽  
Host Organism

What does it take to convert a living organism into a truly productive biofactory? Apart from optimizing biosynthesis pathways as standalone units, a successful bioengineering approach must bend the endogenous metabolic network of the host, and especially its central metabolism, to support the bioproduction process. In practice, this usually involves three complementary strategies which include tuning-down or abolishing competing metabolic pathways, increasing the availability of precursors of the desired biosynthesis pathway, and ensuring high availability of energetic resources such as ATP and NADPH. In this review, we explore these strategies, focusing on key metabolic pathways and processes, such as glycolysis, anaplerosis, the TCA (tricarboxylic acid) cycle, and NADPH production. We show that only a holistic approach for bioengineering — considering the metabolic network of the host organism as a whole, rather than focusing on the production pathway alone — can truly mold microorganisms into efficient biofactories.

scholarly journals Pharmacokinetics, Safety, and Hydrolysis of Oral Pyrroloquinazolinediamines Administered in Single and Multiple Doses in Rats

2007 ◽  
Vol 51 (8) ◽  
pp. 2898-2904 ◽  
Author(s):  
Qigui Li ◽  
Michael P. Kozar ◽  
Todd W. Shearer ◽  
Lisa H. Xie ◽  
Ai J. Lin ◽  
...  
Keyword(s):  
Parent Compound ◽  
Parent Drug ◽  
Therapeutic Index ◽  
High Dose ◽  
Time Curve ◽  
Safety Margins ◽  
Intermediate Metabolites ◽  
Active Intermediate ◽  
Multiple Treatments ◽  
Hydrolysis Of

ABSTRACT Pyrroloquinazolinediamine (PQD) derivatives such as tetra-acetamide PQD (PQD-A4) and bis-ethylcarbamyl PQD (PQD-BE) were much safer (with therapeutic indices of 80 and 32, respectively) than their parent compound, PQD (therapeutic index, 10). Further evaluation of PQD-A4 and PQD-BE in single and multiple pharmacokinetic (PK) studies as well as corresponding toxicity studies was conducted with rats. PQD-A4 could be converted to two intermediate metabolites (monoacetamide PQD and bisacetamide PQD) first and then to the final metabolite, PQD, while PQD-BE was directly hydrolyzed to PQD without precursor and intermediate metabolites. Maximum tolerant doses showed that PQD-A4 and PQD-BE have only 1/12 and 1/6, respectively, of the toxicity of PQD after a single oral dose. Compared to the area under the concentration-time curve for PQD alone (2,965 ng·h/ml), values measured in animals treated with PQD-A4 and PQD-BE were one-third (1,047 ng·h/ml) and one-half (1,381 ng·h/ml) as high, respectively, after an equimolar dosage, suggesting that PQD was the only agent to induce the toxicity. Similar results were also shown in multiple treatments; PQD-A4 and PQD-BE generated two-fifths and three-fifths, respectively, of PQD concentrations, with 8.8-fold and 3.8-fold safety margins, respectively, over the parent drug. PK data indicated that the bioavailability of oral PQD-A4 was greatly limited at high dose levels, that PQD-A4 was slowly converted to PQD via a sequential three-step process of conversion, and that PQD-A4 was significantly less toxic than the one-step hydrolysis drug, PQD-BE. It was concluded that the slow and smaller release of PQD was the main reason for the reduction in toxicity and that the active intermediate metabolites can still maintain antimalarial potency. Therefore, the candidate with multiple-step hydrolysis of PQD could be developed as a safer potential agent for malaria treatment.

scholarly journals Thermal Decomposition Studies of Layered Metal Hydroxynitrates (Metal: Cu, Zn, Cu/Co, and Zn/Co)

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Thimmasandra Narayan Ramesh ◽  
Theeta Lakshamaiah Madhu
Keyword(s):  
Metal Ion ◽  
Structural Transformations ◽  
Single Step ◽  
Cobalt Metal ◽  
Isothermal Decomposition ◽  
Metal Hydroxides ◽  
X Ray ◽  
Metal Nitrates ◽  
Biphasic Mixture ◽  
Hydrolysis Of

Layered metal hydroxynitrates and mixed metal hydroxynitrates (copper/cobalt hydroxynitrates and zinc/cobalt hydroxynitrates at different mole ratios) were synthesized by hydrolysis of urea and metal nitrates at 140°C. Layered metal hydroxyl nitrates derive their structure from brucite mineral and generally crystallize in hexagonal and monoclinic phases. Isothermal decomposition studies of Cu2(OH)3(NO3), Co2(OH)3(NO3), Cu1.5Co0.5(OH)3(NO3), Cu1.34Co0.66(OH)3(NO3), Zn5(OH)8(NO3)2(H2O)2, Zn3.75Co1.25(OH)8(NO3)2(H2O)2, and Zn3.35Co1.65(OH)8(NO3)2(H2O)2 samples were carried out at different intervals of temperature and the structural transformations during the process were monitored using powder X-ray diffractograms. Biphasic mixture of metal hydroxynitrate/metal oxide is observed in case of cobalt/zinc based layered hydroxynitrates, while copper hydroxynitrate or copper/cobalt metal hydroxynitrate decomposes in a single step. The decomposition temperatures of layered metal hydroxynitrates and mixed layered metal hydroxides depend on the method of preparation, their composition and the nature of metal ion, and their coordination.

scholarly journals Highly efficient single-step pretreatment to remove lignin and hemicellulose from softwood

BioResources ◽  
2019 ◽  
Vol 14 (2) ◽  
pp. 3567-3577
Author(s):  
Irma Bernal-Lugo ◽  
Carmen Jacinto-Hernandez ◽  
Miquel Gimeno ◽  
C. Carmina Montiel ◽  
Fausto Rivero-Cruz ◽  
...  
Keyword(s):  
Value Added ◽  
Enzymatic Digestion ◽  
Single Step ◽  
Glucose Yield ◽  
Highly Efficient ◽  
First Case ◽  
Hemicellulose Removal ◽  
One Step ◽  
Low Glucose ◽  
Hydrolysis Of

The use of lignocellulosic softwood residues as feedstock for the production of bioethanol and other value-added chemical products has been limited by its high recalcitrance. Alkaline or organosolvent pretreatments have been used to remove recalcitrance in softwoods. Although these methods partially remove lignin and hemicellulose, they also result in low glucose recovery. In the first case, there is low cellulose hydrolizability, and in the second, there is a loss of cellulose. This study evaluated both methods combined into one step: alkaline hydrolysis of the biomass in the presence of an organosolvent. Different conditions of temperature and residence times were assayed. The efficiency of these conditions was quantified as the percentage of lignin and hemicellulose removed from the biomass without loss of cellulose. The substrate produced with the most efficient conditions removed 91% of the lignin and 89.1% of the hemicellulose with no loss of cellulose. Enzymatic hydrolysis of this biomass was 90% to 95%, with a substrate concentration of 3% and with five filter paper units per gram of cellulose (FPU/g cellulose). These results indicated that this one-step alkaline-organsolvent process, applied as a pretreatment to softwood, allows highly efficient lignin and hemicellulose removal. 100% of cellulose was recovered, and there was between 90 and 95% glucose yield after enzymatic digestion.

Sign in / Sign up

Export Citation Format

Share Document