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.

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 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 A Pathway for Degradation of Uracil to Acetyl Coenzyme A in Bacillus megaterium

2020 ◽  
Vol 86 (7) ◽  
Author(s):  
Di Zhu ◽  
Yifeng Wei ◽  
Jinyu Yin ◽  
Dazhi Liu ◽  
Ee Lui Ang ◽  
...  
Keyword(s):  
Energy Source ◽  
Bacillus Megaterium ◽  
Coenzyme A ◽  
Catabolic Pathway ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Content Type ◽  
Biochemical Pathways ◽  
Acetyl Coenzyme

ABSTRACT Bacteria utilize diverse biochemical pathways for the degradation of the pyrimidine ring. The function of the pathways studied to date has been the release of nitrogen for assimilation. The most widespread of these pathways is the reductive pyrimidine catabolic pathway, which converts uracil into ammonia, carbon dioxide, and β-alanine. Here, we report the characterization of a β-alanine:pyruvate aminotransferase (PydD2) and an NAD+-dependent malonic semialdehyde dehydrogenase (MSDH) from a reductive pyrimidine catabolism gene cluster in Bacillus megaterium. Together, these enzymes convert β-alanine into acetyl coenzyme A (acetyl-CoA), a key intermediate in carbon and energy metabolism. We demonstrate the growth of B. megaterium in defined medium with uracil as its sole carbon and energy source. Homologs of PydD2 and MSDH are found in association with reductive pyrimidine pathway genes in many Gram-positive bacteria in the order Bacillales. Our study provides a basis for further investigations of the utilization of pyrimidines as a carbon and energy source by bacteria. IMPORTANCE Pyrimidine has wide occurrence in natural environments, where bacteria use it as a nitrogen and carbon source for growth. Detailed biochemical pathways have been investigated with focus mainly on nitrogen assimilation in the past decades. Here, we report the discovery and characterization of two important enzymes, PydD2 and MSDH, which constitute an extension for the reductive pyrimidine catabolic pathway. These two enzymes, prevalent in Bacillales based on our bioinformatics studies, allow stepwise conversion of β-alanine, a previous “end product” of the reductive pyrimidine degradation pathway, to acetyl-CoA as carbon and energy source.

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 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 Functional Analyses of Two Acetyl Coenzyme A Synthetases in the Ascomycete Gibberella zeae

2011 ◽  
Vol 10 (8) ◽  
pp. 1043-1052 ◽  
Author(s):  
Seunghoon Lee ◽  
Hokyoung Son ◽  
Jungkwan Lee ◽  
Kyunghun Min ◽  
Gyung Ja Choi ◽  
...  
Keyword(s):  
Sexual Development ◽  
Coenzyme A ◽  
Lipid Synthesis ◽  
Gibberella Zeae ◽  
Atp Citrate Lyase ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Content Type ◽  
Temporal Precision ◽  
Acetyl Coenzyme

ABSTRACTAcetyl coenzyme A (acetyl-CoA) is a crucial metabolite for energy metabolism and biosynthetic pathways and is produced in various cellular compartments with spatial and temporal precision. Our previous study on ATP citrate lyase (ACL) inGibberella zeaerevealed that ACL-dependent acetyl-CoA production is important for histone acetylation, especially in sexual development, but is not involved in lipid synthesis. In this study, we deleted additional acetyl-CoA synthetic genes, the acetyl-CoA synthetases (ACSgenesACS1andACS2), to identify alternative acetyl-CoA production mechanisms for ACL. TheACS1deletion resulted in a defect in sexual development that was mainly due to a reduction in 1-palmitoyl-2-oleoyl-3-linoleoyl-rac-glycerol production, which is required for perithecium development and maturation. Another ACS coding gene,ACS2, has accessorial functions forACS1and has compensatory functions forACLas a nuclear acetyl-CoA producer. This study showed that acetate is readily generated during the entire life cycle ofG. zeaeand has a pivotal role in fungal metabolism. Because ACSs are components of the pyruvate-acetaldehyde-acetate pathway, this fermentation process might have crucial roles in various physiological processes for filamentous fungi.

scholarly journals Synthetic Biology for Engineering Acetyl Coenzyme A Metabolism in Yeast

mBio ◽  
2014 ◽  
Vol 5 (6) ◽  
Author(s):  
Jens Nielsen
Keyword(s):  
Coenzyme A ◽  
Pyruvate Dehydrogenase Complex ◽  
High Yield ◽  
Cell Factory ◽  
Efficient Production ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Content Type ◽  
Acetyl Coenzyme ◽  
High Yields

ABSTRACT The yeast Saccharomyces cerevisiae is a widely used cell factory for the production of fuels, chemicals, and pharmaceuticals. The use of this cell factory for cost-efficient production of novel fuels and chemicals requires high yields and low by-product production. Many industrially interesting chemicals are biosynthesized from acetyl coenzyme A (acetyl-CoA), which serves as a central precursor metabolite in yeast. To ensure high yields in production of these chemicals, it is necessary to engineer the central carbon metabolism so that ethanol production is minimized (or eliminated) and acetyl-CoA can be formed from glucose in high yield. Here the perspective of generating yeast platform strains that have such properties is discussed in the context of a major breakthrough with expression of a functional pyruvate dehydrogenase complex in the cytosol.

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 FINE STRUCTURAL LOCALIZATION OF ACETYL COENZYME A CARBOXYLASE IN RAT HEPATOCYTES

1969 ◽  
Vol 17 (6) ◽  
pp. 379-385 ◽  
Author(s):  
R. D. YATES ◽  
JOAN A. HIGGINS ◽  
H. J. BARRNETT
Keyword(s):  
Fatty Acid Biosynthesis ◽  
Coenzyme A ◽  
Rat Hepatocytes ◽  
Adenosine Diphosphate ◽  
Complete Medium ◽  
Lead Phosphate ◽  
Acetyl Coa ◽  
Structural Localization ◽  
Acetyl Coenzyme A ◽  
Acetyl Coenzyme

Acetyl coenzyme A (CoA) carboxylase, the enzyme catalyzing the first step in fatty acid biosynthesis, has been localized in the hepatocytes of glutaraldehyde-fixed rat livers. This reaction in which the hydrolysis of adenosine triphosphate (ATP) is coupled to the synthesis of malonyl-CoA utilizes biotin, bicarbonate, acetyl-CoA, ATP and manganese or magnesium. ATP hydrolysis yields adenosine diphosphate and phosphate, the latter of which is subsequently converted to a lead phosphate precipitate. In tissues incubated in a complete medium, lead phosphate was seen to be closely associated with the outer surfaces of the membranes of the granular endoplasmic reticulum. When ATP was omitted from the medium no intracellular lead deposit was noted. A small amount of reaction product was seen when bicarbonate, biotin or acetyl-CoA was omitted from the medium. Reaction product was not seen when the biotin inhibitor avidin was added to the incubation medium or when tissue sections were boiled in buffer prior to incubation. The possibility of using a similar technique in studying other coupled enzyme systems is considered.

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