scholarly journals Presence of Acetyl Coenzyme A (CoA) Carboxylase and Propionyl-CoA Carboxylase in Autotrophic Crenarchaeota and Indication for Operation of a 3-Hydroxypropionate Cycle in Autotrophic Carbon Fixation

1999 ◽  
Vol 181 (4) ◽  
pp. 1088-1098 ◽  
Author(s):  
Castor Menendez ◽  
Zsuzsa Bauer ◽  
Harald Huber ◽  
Nasser Gad’on ◽  
Karl-Otto Stetter ◽  
...  
Keyword(s):  
Phosphoenolpyruvate Carboxylase ◽  
Carbon Fixation ◽  
Coenzyme A ◽  
Autotrophic Growth ◽  
Acetyl Coa Carboxylase ◽  
Acetyl Coa ◽  
Carboxylase Activity ◽  
Acetyl Coenzyme A ◽  
Acetyl Coenzyme

ABSTRACT The pathway of autotrophic CO2 fixation was studied in the phototrophic bacterium Chloroflexus aurantiacus and in the aerobic thermoacidophilic archaeon Metallosphaera sedula. In both organisms, none of the key enzymes of the reductive pentose phosphate cycle, the reductive citric acid cycle, and the reductive acetyl coenzyme A (acetyl-CoA) pathway were detectable. However, cells contained the biotin-dependent acetyl-CoA carboxylase and propionyl-CoA carboxylase as well as phosphoenolpyruvate carboxylase. The specific enzyme activities of the carboxylases were high enough to explain the autotrophic growth rate via the 3-hydroxypropionate cycle. Extracts catalyzed the CO2-, MgATP-, and NADPH-dependent conversion of acetyl-CoA to 3-hydroxypropionate via malonyl-CoA and the conversion of this intermediate to succinate via propionyl-CoA. The labelled intermediates were detected in vitro with either 14CO2 or [14C]acetyl-CoA as precursor. These reactions are part of the 3-hydroxypropionate cycle, the autotrophic pathway proposed forC. aurantiacus. The investigation was extended to the autotrophic archaea Sulfolobus metallicus andAcidianus infernus, which showed acetyl-CoA and propionyl-CoA carboxylase activities in extracts of autotrophically grown cells. Acetyl-CoA carboxylase activity is unexpected in archaea since they do not contain fatty acids in their membranes. These aerobic archaea, as well as C. aurantiacus, were screened for biotin-containing proteins by the avidin-peroxidase test. They contained large amounts of a small biotin-carrying protein, which is most likely part of the acetyl-CoA and propionyl-CoA carboxylases. Other archaea reported to use one of the other known autotrophic pathways lacked such small biotin-containing proteins. These findings suggest that the aerobic autotrophic archaea M. sedula,S. metallicus, and A. infernus use a yet-to-be-defined 3-hydroxypropionate cycle for their autotrophic growth. Acetyl-CoA carboxylase and propionyl-CoA carboxylase are proposed to be the main CO2 fixation enzymes, and phosphoenolpyruvate carboxylase may have an anaplerotic function. The results also provide further support for the occurrence of the 3-hydroxypropionate cycle in C. aurantiacus.

scholarly journals Genome Sequence of Serratia marcescens MSU97, a Plant-Associated Bacterium That Makes Multiple Antibiotics

2017 ◽  
Vol 5 (9) ◽  
Author(s):  
Miguel A. Matilla ◽  
Zulema Udaondo ◽  
Tino Krell ◽  
George P. C. Salmond
Keyword(s):  
Serratia Marcescens ◽  
Genome Sequence ◽  
Coenzyme A ◽  
Acetyl Coa Carboxylase ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Content Type ◽  
Acetyl Coenzyme ◽  
Multiple Antibiotics

ABSTRACT Serratia marcescens MSU97 was isolated from the Guayana region of Venezuela due to its ability to suppress plant-pathogenic oomycetes. Here, we report the genome sequence of MSU97, which produces various antibiotics, including the bacterial acetyl-coenzyme A (acetyl-CoA) carboxylase inhibitor andrimid, the chlorinated macrolide oocydin A, and the red linear tripyrrole antibiotic prodigiosin.

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 Isolated Poly(3-Hydroxybutyrate) (PHB) Granules Are Complex Bacterial Organelles Catalyzing Formation of PHB from Acetyl Coenzyme A (CoA) and Degradation of PHB to Acetyl-CoA

2007 ◽  
Vol 189 (22) ◽  
pp. 8250-8256 ◽  
Author(s):  
Keiichi Uchino ◽  
Terumi Saito ◽  
Birgit Gebauer ◽  
Dieter Jendrossek
Keyword(s):  
Coenzyme A ◽  
Ralstonia Eutropha ◽  
Major Product ◽  
Acetyl Coa ◽  
Native Form ◽  
Acetyl Coenzyme A ◽  
Acetyl Coenzyme ◽  
Phb Synthase ◽  
Coa Reductase

ABSTRACTPoly(3-hydroxybutyrate) (PHB) granules isolated in native form (nPHB granules) fromRalstonia eutrophacatalyzed formation of PHB from14C-labeled acetyl coenzyme A (CoA) in the presence of NADPH and concomitantly released CoA, revealing that PHB biosynthetic proteins (acetoacetyl-CoA thiolase, acetoacetyl-CoA reductase, and PHB synthase) are present and active in isolated nPHB granules in vitro. nPHB granules also catalyzed thiolytic cleavage of PHB in the presence of added CoA, resulting in synthesis of 3-hydroxybutyryl-CoA (3HB-CoA) from PHB. Synthesis of 3HB-CoA was also shown by incubation of artificial (protein-free) PHB with CoA and PhaZa1, confirming that PhaZa1 is a PHB depolymerase catalyzing the thiolysis reaction. Acetyl-CoA was the major product detectable after incubation of nPHB granules in the presence of NAD+, indicating that downstream mobilizing enzyme activities were also present and active in isolated nPHB granules. We propose that intracellular concentrations of key metabolites (CoA, acetyl-CoA, 3HB-CoA, NAD+/NADH) determine whether a cell accumulates or degrades PHB. Since the degradation product of PHB is 3HB-CoA, the cells do not waste energy by synthesis and degradation of PHB. Thus, our results explain the frequent finding of simultaneous synthesis and breakdown of PHB.

scholarly journals Activation of immobilized acetyl-Coenzyme A carboxylase

1978 ◽  
Vol 169 (1) ◽  
pp. 255-256 ◽  
Author(s):  
A D Landman ◽  
J Lampert
Keyword(s):  
Coenzyme A ◽  
Enzymic Activity ◽  
Acetyl Coa Carboxylase ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Acetyl Coenzyme ◽  
Sepharose 4B ◽  
Acetyl Coenzyme A Carboxylase ◽  
Covalently Bound

Partially purified acetyl-CoA carboxylase was covalently bound to a Sepharose 4B matrix. Although aggregation was thus prevented, the enzymic activity was stimulated by citrate and isocitrate.

scholarly journals Investigation of Carbon Metabolism in “Dehalococcoides ethenogenes” Strain 195 by Use of Isotopomer and Transcriptomic Analyses

2009 ◽  
Vol 191 (16) ◽  
pp. 5224-5231 ◽  
Author(s):  
Yinjie J. Tang ◽  
Shan Yi ◽  
Wei-Qin Zhuang ◽  
Stephen H. Zinder ◽  
Jay D. Keasling ◽  
...  
Keyword(s):  
Genome Annotation ◽  
Carbon Fixation ◽  
Coenzyme A ◽  
Citrate Synthase ◽  
Tricarboxylic Acid Cycle ◽  
Acetyl Coa ◽  
Experimental Conditions ◽  
Acetyl Coenzyme A ◽  
Acetyl Coenzyme ◽  
Dehalococcoides Ethenogenes

ABSTRACT Members of the genus “Dehalococcoides” are the only known microorganisms that can completely dechlorinate tetrachloroethene and trichloroethene to the innocuous end product, ethene. This study examines the central metabolism in “Dehalococcoides ethenogenes” strain 195 via 13C-labeled tracer experiments. Supported by the genome annotation and the transcript profile, isotopomer analysis of key metabolites clarifies ambiguities in the genome annotation and identifies an unusual biosynthetic pathway in strain 195. First, the 13C-labeling studies revealed that strain 195 contains complete amino acid biosynthesis pathways, even though current genome annotation suggests that several of these pathways are incomplete. Second, the tricarboxylic acid cycle of strain 195 is confirmed to be branched, and the Wood-Ljungdahl carbon fixation pathway is shown to not be functionally active under our experimental conditions; rather, CO2 is assimilated via two reactions, conversion of acetyl-coenzyme A (acetyl coenzyme A [acetyl-CoA]) to pyruvate catalyzed by pyruvate synthase (DET0724-0727) and pyruvate conversion to oxaloacetate via pyruvate carboxylase (DET0119-0120). Third, the 13C-labeling studies also suggested that isoleucine is synthesized from acetyl-CoA and pyruvate via citramalate synthase (CimA, EC 2.3.1.182), rather than from the common pathway via threonine ammonia-lyase (EC 4.3.1.19). Finally, evidence is presented that strain 195 may contain an undocumented citrate synthase (>95% Re-type stereospecific), i.e., a novel Re-citrate synthase that is apparently different from the one recently reported in Clostridium kluyveri.

scholarly journals PanM, an Acetyl-Coenzyme A Sensor Required for Maturation of l-Aspartate Decarboxylase (PanD)

mBio ◽  
2012 ◽  
Vol 3 (4) ◽  
Author(s):  
Tara N. Stuecker ◽  
Alex C. Tucker ◽  
Jorge C. Escalante-Semerena
Keyword(s):  
Coenzyme A ◽  
Lysine Acetylation ◽  
Acetyl Coa ◽  
Transfer Event ◽  
Acetyl Coenzyme A ◽  
Content Type ◽  
Acetyltransferase Activity ◽  
Acetyl Coenzyme

ABSTRACTCoenzyme A (CoA) is essential for cellular chemistry in all forms of life. The pantothenate moiety of CoA is generated from the condensation of pantoate and β-alanine. β-Alanine is formed by decarboxylation ofl-aspartate catalyzed by PanD, a pyruvoyl enzyme that is synthesized by the cell as an inactive precursor (pro-PanD). Maturation of pro-PanD into PanD occurs via a self-cleavage event at residue Ser25, which forms the catalytic pyruvoyl moiety. We recently reported thatSalmonella entericaPanM was necessary for pro-PanD maturation, bothin vitroandin vivo. Notably, PanM is annotated as a Gcn5-likeN-acetyltransferase (GNAT), which suggested that lysine acetylation might be part of the mechanism of maturation. Here we show that PanM lacks acetyltransferase activity and that acetyl-CoA stimulates its activity. Results of experiments with nonhydrolyzable ethyl-CoA and genetically encoded acetyl-lysine-containing PanD support the conclusion that PanM-dependent pro-PanD maturation does not involve an acetyl transfer event. We also show that CoA binding to PanM is needed forin vivoactivity and that disruption of CoA binding prevents PanM from interacting with PanD. We conclude that PanM is a GNAT homologue that lost its acetyltransferase activity and evolved a new function as an acetyl-CoA sensor that can trigger the maturation of pro-PanD.IMPORTANCENε-lysine acetylation is increasingly being recognized as a widespread and important form of posttranslational regulation in bacteria. The acetyltransferases that catalyze these reactions are poorly characterized in bacteria. Based on annotation, most bacterial genomes contain several acetyltransferases, but the physiological roles of only a handful have been determined. Notably, a subset of putative acetyltransferases lack residues that are critical for activity in most biochemically characterized acetyltransferases. We show that one such putative acetyltransferase, PanM (formerly YhhK), lacks acetyltransferase activity but functions instead as an acetyl-coenzyme A (CoA) sensor. This work establishes the possibility that, like PanM, other putative acetyltransferases may have evolved new functions while retaining the ability to sense acetyl-CoA.

scholarly journals Novel Bacterial Acetyl Coenzyme A Carboxylase Inhibitors with Antibiotic Efficacy In Vivo

2006 ◽  
Vol 50 (8) ◽  
pp. 2707-2712 ◽  
Author(s):  
C. Freiberg ◽  
J. Pohlmann ◽  
P. G. Nell ◽  
R. Endermann ◽  
J. Schuhmacher ◽  
...  
Keyword(s):  
Coenzyme A ◽  
Cross Resistance ◽  
Resistance Mutations ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
In Vivo Efficacy ◽  
Acetyl Coenzyme ◽  
Significant Efficacy

ABSTRACT The pseudopeptide pyrrolidinedione antibiotics, such as moiramide B, have recently been discovered to target the multisubunit acetyl coenzyme A (acetyl-CoA) carboxylases of bacteria. In this paper, we describe synthetic variations of each moiety of the modularly composed pyrrolidinediones, providing insight into structure-activity relationships of biochemical target activity, in vitro potency, and in vivo efficacy. The novel derivatives showed highly improved activities against gram-positive bacteria compared to those of previously reported variants. The compounds exhibited a MIC90 value of 0.1 μg/ml against a broad spectrum of Staphylococcus aureus clinical isolates. No cross-resistance to antibiotics currently used in clinical practice was observed. Resistance mutations induced by pyrrolidinediones are exclusively located in the carboxyltransferase subunits of the bacterial acetyl-CoA carboxylase, indicating the identical mechanisms of action of all derivatives tested. Improvement of the physicochemical profile was achieved by salt formation, leading to aqueous solubilities of up to 5 g/liter. For the first time, the in vitro activity of this compound class was compared with its in vivo efficacy, demonstrating a path from compounds weakly active in vivo to agents with significant efficacy. In a murine model of S. aureus sepsis, the 100% effective dose of the best compound reported was 25 mg/kg of body weight, only fourfold higher than that of the comparator molecule linezolid. The obvious improvements achieved by chemical derivatization reflect the potential of this novel antibiotic compound class for future therapy.

scholarly journals Deconstructing Methanosarcina acetivorans into an acetogenic archaeon

2022 ◽  
Vol 119 (2) ◽  
pp. e2113853119
Author(s):  
Christian Schöne ◽  
Anja Poehlein ◽  
Nico Jehmlich ◽  
Norman Adlung ◽  
Rolf Daniel ◽  
...  
Keyword(s):  
Carbon Monoxide ◽  
Energy Conservation ◽  
Carbon Fixation ◽  
Coenzyme A ◽  
Methanogenic Archaea ◽  
Methanosarcina Acetivorans ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Acetyl Coenzyme ◽  
Acetyl Coa Pathway

The reductive acetyl-coenzyme A (acetyl-CoA) pathway, whereby carbon dioxide is sequentially reduced to acetyl-CoA via coenzyme-bound C1 intermediates, is the only autotrophic pathway that can at the same time be the means for energy conservation. A conceptually similar metabolism and a key process in the global carbon cycle is methanogenesis, the biogenic formation of methane. All known methanogenic archaea depend on methanogenesis to sustain growth and use the reductive acetyl-CoA pathway for autotrophic carbon fixation. Here, we converted a methanogen into an acetogen and show that Methanosarcina acetivorans can dispense with methanogenesis for energy conservation completely. By targeted disruption of the methanogenic pathway, followed by adaptive evolution, a strain was created that sustained growth via carbon monoxide–dependent acetogenesis. A minute flux (less than 0.2% of the carbon monoxide consumed) through the methane-liberating reaction remained essential, indicating that currently living methanogens utilize metabolites of this reaction also for anabolic purposes. These results suggest that the metabolic flexibility of methanogenic archaea might be much greater than currently known. Also, our ability to deconstruct a methanogen into an acetogen by merely removing cellular functions provides experimental support for the notion that methanogenesis could have evolved from the reductive acetyl-coenzyme A pathway.

scholarly journals Acetyl Coenzyme A Synthetase (ADP Forming) from the Hyperthermophilic Archaeon Pyrococcus furiosus: Identification, Cloning, Separate Expression of the Encoding Genes,acdAI and acdBI, in Escherichia coli, and In Vitro Reconstitution of the Active Heterotetrameric Enzyme from Its Recombinant Subunits

1999 ◽  
Vol 181 (18) ◽  
pp. 5885-5888 ◽  
Author(s):  
Meike Musfeldt ◽  
Martina Selig ◽  
Peter Schönheit
Keyword(s):  
Escherichia Coli ◽  
Coenzyme A ◽  
Pyrococcus Furiosus ◽  
Amino Acid Sequences ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Acetyl Coenzyme ◽  
Acetate Formation ◽  
Encoding Genes

ABSTRACT Acetyl-coenzyme A (acetyl-CoA) synthetase (ADP forming) represents a novel enzyme in archaea of acetate formation and energy conservation (acetyl-CoA + ADP + Pi → acetate + ATP + CoA). Two isoforms of the enzyme have been purified from the hyperthermophile Pyrococcus furiosus. Isoform I is a heterotetramer (α2β2) with an apparent molecular mass of 145 kDa, composed of two subunits, α and β, with apparent molecular masses of 47 and 25 kDa, respectively. By using N-terminal amino acid sequences of both subunits, the encoding genes, designated acdAI and acdBI, were identified in the genome of P. furiosus. The genes were separately overexpressed in Escherichia coli, and the recombinant subunits were reconstituted in vitro to the active heterotetrameric enzyme. The purified recombinant enzyme showed molecular and catalytical properties very similar to those shown by acetyl-CoA synthetase (ADP forming) purified from P. furiosus.

scholarly journals Structure, Activity, and Inhibition of the Carboxyltransferase β-Subunit of Acetyl Coenzyme A Carboxylase (AccD6) from Mycobacterium tuberculosis

2014 ◽  
Vol 58 (10) ◽  
pp. 6122-6132 ◽  
Author(s):  
Manchi C. M. Reddy ◽  
Ardala Breda ◽  
John B. Bruning ◽  
Mukul Sherekar ◽  
Spandana Valluru ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Fatty Acid Biosynthesis ◽  
Coenzyme A ◽  
Cell Activity ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Content Type ◽  
Acetyl Coenzyme ◽  
Malonyl Coa

ABSTRACTInMycobacterium tuberculosis, the carboxylation of acetyl coenzyme A (acetyl-CoA) to produce malonyl-CoA, a building block in long-chain fatty acid biosynthesis, is catalyzed by two enzymes working sequentially: a biotin carboxylase (AccA) and a carboxyltransferase (AccD). While the exact roles of the three different biotin carboxylases (AccA1 to -3) and the six carboxyltransferases (AccD1 to -6) inM. tuberculosisare still not clear, AccD6 in complex with AccA3 can synthesize malonyl-CoA from acetyl-CoA. A series of 10 herbicides that target plant acetyl-CoA carboxylases (ACC) were tested for inhibition of AccD6 and for whole-cell activity againstM. tuberculosis. From the tested herbicides, haloxyfop, an arylophenoxypropionate, showedin vitroinhibition ofM. tuberculosisAccD6, with a 50% inhibitory concentration (IC50) of 21.4 ± 1 μM. Here, we report the crystal structures ofM. tuberculosisAccD6 in the apo form (3.0 Å) and in complex with haloxyfop-R(2.3 Å). The structure ofM. tuberculosisAccD6 in complex with haloxyfop-Rshows two molecules of the inhibitor bound on each AccD6 subunit. These results indicate the potential for developing novel therapeutics for tuberculosis based on herbicides with low human toxicity.

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