Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:6.2.1.13 (acetyl-CoA synthetase)
451 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

High rates of methanogenesis from acetate and ATP were observed from cell-free extracts of the thermophilic acetotrophic methanogen Methanothrix (Methanosaeta) thermophila strain CALS-1 when cultures were grown in a pH auxostat fed with acetic acid. Specific methanogenic activities ranged from 50-300 nmol min-1 (mg protein)-1, which was comparable to those for whole cells. In contrast to results with Methanosarcina spp., the reaction did not require high levels of H2 in the headspace. CO was inhibitory to methanogenesis from acetate. The inhibition by CO and the lack of effect of H2 on methanogenesis from acetate resemble previous results with whole cells of CALS-1. Protein concentrations in extracts > 5 mg/ml were required for good activity, and the optimum temperature for the methanogenesis was near 65° C. ATP was required in substrate quantities and was converted mainly to AMP. The maximum CH4/ATP stoichiometry obtained was near 1.0, consistent with acetate activation using an acetyl-CoA synthetase mechanism that converts ATP to AMP and pyrophosphate. Methanogenesis in extracts was inhibited by bromoethane sulfonate and cyanide, indicating the involvement of methylcoenzyme M methylreductase and a carbon monoxide dehydrogenase complex with methanogenesis from acetate. These results are consistent with acetyl-coenzyme A (CoA) as the form of activated acetate involved in methanogenesis from acetate in strain CALS-1, but no activity could be obtained from extracts using acetyl-CoA as a substrate.
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PMID:Methanogenesis from acetate by cell-free extracts of the thermophilic acetotrophic methanogen Methanothrix thermophila CALS-1 882 51

The filamentous fungus Aspergillus nidulans is able to grow on phenylacetic acid (PhAc) as the sole carbon source and has a highly specific phenylacetic acid transport system mediating the uptake of this aromatic compound. This transport system is also able to transport some phenoxyacetic acid (PhOAc), although less efficiently. Maximal uptake rates were observed at 37 degrees C in 50 mM phosphate buffer (pH 7.0). Under these conditions, uptake was linear for at least 1 minute, with K(m) values for PhAc and PhOAc of 74 and 425 microM, respectively. The PhAc transport system is strongly induced by PhAc and, to a lesser extent by PhOAc and other phenyl derivatives. The utilization of glucose (and other sugars), glycerol or acetate results in a substantially reduced uptake. This negative effect caused by certain carbon sources is independent of the creA gene, the regulatory gene mediating carbon catabolite repression. Negative regulation by acetate is prevented by a loss-of-function mutation in the gene encoding acetyl-CoA synthetase, strongly suggesting that this regulation is mediated by the intracellular pool of acetyl-CoA.
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PMID:The phenylacetic acid uptake system of Aspergillus nidulans is under a creA-independent model of catabolic repression which seems to be mediated by acetyl-CoA. 906 65

Acetyl-CoA synthetase (ADP-forming) is an enzyme in Archaea that catalyzes the formation of acetate from acetyl-CoA and couples this reaction with the synthesis of ATP from ADP and Pi (acetyl-CoA + ADP + Pi --> acetate + ATP + CoA) [Schifer, T., Selig, M. & Schonheit, P. (1993) Arch. Microbiol. 159, 72-83]. The enzyme from the anaerobic hyperthermophile Pyrococcus furiosus was purified 96-fold with a yield of 20% to apparent electrophoretic homogeneity. The oxygen-stable enzyme had an apparent molecular mass of 145 kDa and was composed of two subunits with apparent molecular masses of 47 kDa and 25 kDa, indicating an alpha2beta2 structure. The N-terminal amino acid sequences of both subunits were determined; they do not show significant identity to other proteins in databases. The purified enzyme catalyzed the reversible conversion of acetyl-CoA, ADP and Pi to acetate, ATP and CoA. The apparent Vmax value in the direction of acetate formation was 18 U/mg (55 degrees C), the apparent Km values for acetyl-CoA, ADP and Pi were 17 microM, 60 microM and 200 microM, respectively. ADP and Pi could not be replaced by AMP and PPi, defining the enzyme as an ADP-forming rather than an AMP-forming acetyl-CoA synthetase. The apparent Vmax value in the direction of acetyl-CoA formation was about 40 U/mg (55 degrees C), and the apparent Km values for acetate, ATP and CoA were 660 microM, 80 microM and 30 microM, respectively. The purified enzyme was not specific for acetyl-CoA or acetate, in addition to acetyl-CoA (100%), the enzyme accepts propionyl-CoA (110%) and butyryl-CoA (92%), and in addition to acetate (100%), the enzyme accepts propionate (100%), butyrate (92%), isobutyrate (79%), valerate (36%) and isovalerate (34%), indicating that the enzyme functions as an acyl-CoA synthetase (ADP-forming) with a broad substrate spectrum. Succinate, phenylacetate and indoleacetate did not serve as substrates for the enzyme (<3%). In addition to ADP (100%), GDP (220%) and IDP (250%) were used, and in addition to ATP (100%), GTP (210%) and ITP (320%) were used. Pyrimidine nucleotides were not accepted. The enzyme was dependent on Mg2+, which could be partly substituted by Mn2+ and Co2+. The pH optimum was pH 7. The enzyme has a temperature optimum at 90 degrees C, which is in accordance with its physiological function under hyperthermophilic conditions. The enzyme was stabilized against heat inactivation by salts. In the presence of KCI (1 M), which was most effective, the enzyme did not loose activity after 2 h incubation at 100 degrees C.
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PMID:Purification and properties of acetyl-CoA synthetase (ADP-forming), an archaeal enzyme of acetate formation and ATP synthesis, from the hyperthermophile Pyrococcus furiosus. 911 24

The yeast Saccharomyces cerevisiae contains two acetyl-CoA synthetase genes, ACS1 and ACS2. While ACS1 transcription is glucose repressible, ACS2 shows coregulation with structural genes of fatty acid biosynthesis. The ACS2 upstream region contains an ICRE (inositol/choline-responsive element) as an activating sequence and requires the regulatory genes INO2 and INO4 for maximal expression. We demonstrate in vitro binding of the heterodimeric activator protein Ino2p/Ino4p to the ACS2 promoter. In addition, the pleiotropic transcription factor Abf1p also binds to the ACS2 control region. The identification of ACS2 activating elements also found upstream of ACC1, FAS1 and FAS2 suggests a role of this acetyl-CoA synthetase isoenzyme for the generation of the acetyl-CoA pool required for fatty acid biosynthesis.
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PMID:The acetyl-CoA synthetase gene ACS2 of the yeast Saccharomyces cerevisiae is coregulated with structural genes of fatty acid biosynthesis by the transcriptional activators Ino2p and Ino4p. 932 60

Acetaldehyde is one of the intermediate products of ethanolic fermentation, which can be reduced to ethanol by alcohol dehydrogenase (ADH). Alternatively, acetaldehyde can be oxidized to acetate by aldehyde dehydrogenase (ALDH) and subsequently converted to acetyl-CoA by acetyl-CoA synthetase (ACS). To study the expression of ALDHs in plants we isolated and characterized a cDNA coding for a putative mitochondrial ALDH (TobAldh2A) in Nicotiana tabacum. TobALDH2A shows 54-60% identity at the amino acid level with other ALDHs and shows 76% identity with maize Rf2, a gene involved in restoration of male fertility in cms-T maize. TobAldh2A transcripts and protein were present at high levels in the male and female reproductive tissues. Expression in vegetative tissues was much lower and no induction by anaerobic incubation was observed. This suggests that TobALDH expression is not part of the anaerobic response, but may have another function. The use of specific inhibitors of ALDH and the pyruvate dehydrogenase (PDH) complex indicates that ALDH activity is important for pollen tube growth, and thus may have a function in biosynthesis or energy production.
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PMID:Aldehyde dehydrogenase in tobacco pollen. 934 59

Concentrations of total CoAs in chloroplasts freshly isolated from spinach and peas were 10-20 microM, assuming a stromal volume of 66 microl per mg of chlorophyll. Acetyl-CoA and CoASH constituted at least 90% of the total CoA in freshly isolated chloroplasts. For a given chloroplast preparation, the concentration of endogenous acetyl-CoA was the same when extractions were performed using HClO4, trichloroacetic acid, propan-2-ol or chloroform/methanol, and the extracts analysed by quantitative HPLC after minimal processing. During fatty acid synthesis from acetate, concentrations of CoASH within spinach and pea chloroplasts varied from less than 0.1 to 5.0 microM. Malonyl-CoA concentrations were also very low (<0.1-3.0 microM) during fatty acid synthesis but could be calculated from radioactivity incorporated from [1-14C]acetate. Concentrations of CoASH in chloroplasts synthesizing fatty acids could be doubled in the presence of Triton X-100, suggesting that the detergent stimulates fatty acid synthesis by increasing the turnover rate of acyl-CoA. However, although taken up, exogenous CoASH (1 microM) did not stimulate fatty acid synthesis by permeabilized spinach chloroplasts. Calculated rates for acetyl-CoA synthetase, acetyl-CoA carboxylase and malonyl-CoA-acyl-carrier protein transacylase reactions at the concentrations of metabolites measured here are < 0.1-4% of the observed rates of fatty acid synthesis from acetate by isolated chloroplasts. The results suggest that CoA and its esters are probably confined within, and channelled through, the initial stages of a fatty acid synthase multienzyme complex.
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PMID:Stromal concentrations of coenzyme A and its esters are insufficient to account for rates of chloroplast fatty acid synthesis: evidence for substrate channelling within the chloroplast fatty acid synthase. 935 62

The KlPDA1 gene, encoding the E1alpha subunit of the mitochondrial pyruvate-dehydrogenase (PDH) complex was isolated from a Kluyveromyces lactis genomic library by screening with a 1.1 kb internal fragment of the Saccharomyces cerevisiae PDA1 gene. The predicted amino acid sequence encoded by KlPDA1 showed 87% similarity and 79% identity to its S. cerevisiae counterpart. Disruption of KIPDA1 resulted in complete absence of PDH activity in cell extracts. The maximum specific growth rate on glucose of null mutants was 3.5-fold lower than that of the wild-type, whereas growth on ethanol was unaffected. Wild-type K. lactis CBS 2359 exhibits a Crabtree-negative phenotype, i.e. no ethanol was produced in aerobic batch cultures grown on glucose. In contrast, substantial amounts of ethanol and acetaldehyde were produced in aerobic cultures of an isogenic Klpda1 null mutant. A wild-type specific growth rate was restored after introduction of an intact KlPDA1 gene but not, as previously found for S. cerevisiae pda1 mutants, by cultivation in the presence of leucine. The occurrence of aerobic fermentation and slow growth of the Klpda1 null mutant indicate that, although present, the enzymes of the PDH bypass (pyruvate decarboxylase, acetaldehyde dehydrogenase and acetyl-CoA synthetase) could not efficiently replace the PDH complex during batch cultivation on glucose. Only at relatively low growth rates (D = 0.10 h(-1)) in aerobic, glucose-limited chemostat cultures, could the PDH bypass completely replace the PDH complex, thus allowing fully respiratory growth. This resulted in a lower biomass yield [g biomass (g glucose)-1] than in the wild-type due to a higher consumption of ATP in the PDH bypass compared to the formation of acetyl-CoA via the PDH complex.
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PMID:Inactivation of the Kluyveromyces lactis KlPDA1 gene leads to loss of pyruvate dehydrogenase activity, impairs growth on glucose and triggers aerobic alcoholic fermentation. 988 36

To identify transcriptionally regulated mediators associated with the cell cycle, we adopted the differential mRNA display technique for cell cultures of Tetrahymena pyriformis synchronized by cyclic heat treatment. One cDNA fragment that was expressed differently during synchronous cell division had a greatly decreased expression at 30 min after the end of heat treatment (EHT). Using this fragment as a probe, we isolated the full-length cDNA for T. pyriformis acetyl-CoA synthetase (TpAcs) which encodes a 651 amino acid polypeptide with a predicted molecular mass of 72.8 kDa. The deduced amino acid sequence of T. pyriformis ACS shows 42% sequence identity compared with that of Lysobacter sp. acetyl-CoA synthetase (ACS), an enzyme which catalyses the formation of acetyl-CoA from acetate via an acetyl-adenylate intermediate. The deduced sequence is also 41% and 40% identical compared with those of Pseudomonas putida and Coprinus cinereus ACS, respectively. The deduced sequence of T. pyriformis ACS also shares similar characteristics of the conserved motifs I and II in the ACS family. To further investigate the actions of the gene encoding this enzyme, mRNA expression was determined during the course of synchronized cell division in T. pyriformis. Northern blot results show that the mRNA level was dramatically decreased at 30 min after EHT prior to entering synchronous cell division (which occurs 75 min after EHT), suggesting that mRNA expression of the TpAcs was associated with the cell cycle and that the down-regulated expression of TpAcs at 30 min after EHT would be required for the initiation of the oncoming synchronous cell division in T. pyriformis.
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PMID:Molecular cloning and cell-cycle-dependent expression of the acetyl-CoA synthetase gene in Tetrahymena cells. 1051 Mar 17

The gene coding for the acetyl-CoA synthetase (ADP-forming) from the amitochondriate eukaryote Giardia lamblia has been expressed in Escherichia coli. The recombinant enzyme exhibited the same substrate specificity as the native enzyme, utilizing acetyl-CoA and adenine nucleotides as preferred substrates and less efficiently, propionyl- and succinyl-CoA. N- and C-terminal parts of the G. lamblia acetyl-CoA synthetase sequence were found to be homologous to the alpha- and beta-subunits, respectively, of succinyl-CoA synthetase. Sequence analysis of homologous enzymes from various bacteria, archaea, and the eukaryote, Plasmodium falciparum, identified conserved features in their organization, which allowed us to delineate a new superfamily of acyl-CoA synthetases (nucleoside diphosphate-forming) and its signature motifs. The representatives of this new superfamily of thiokinases vary in their domain arrangement, some consisting of separate alpha- and beta-subunits and others comprising fusion proteins in alpha-beta or beta-alpha orientation. The presence of homologs of acetyl-CoA synthetase (ADP-forming) in such human pathogens as G. lamblia, Yersinia pestis, Bordetella pertussis, Pseudomonas aeruginosa, Vibrio cholerae, Salmonella typhi, Porphyromonas gingivalis, and the malaria agent P. falciparum suggests that they might be used as potential drug targets.
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PMID:Acetyl-CoA synthetase from the amitochondriate eukaryote Giardia lamblia belongs to the newly recognized superfamily of acyl-CoA synthetases (Nucleoside diphosphate-forming). 1068 68

Through suppressive subtractive hybridization, we identified a new gene whose transcription is induced by sterol regulatory element-binding proteins (SREBPs). The gene encodes acetyl-CoA synthetase (ACS), the cytosolic enzyme that activates acetate so that it can be used for lipid synthesis or for energy generation. ACS genes were isolated previously from yeast, but not from animal cells. Recombinant human ACS was produced by expressing the cloned cDNA transiently in human cells. After purification by nickel chromatography, the 701-amino acid cytosolic enzyme was shown to function as a monomer. The recombinant enzyme produced acetyl-CoA from acetate in a reaction that required ATP. As expected for a gene controlled by SREBPs, ACS mRNA was induced when cultured cells were deprived of sterols and repressed by sterol addition. The pattern of regulation resembled the regulation of enzymes of fatty acid synthesis. ACS mRNA was also elevated in livers of transgenic mice that express dominant-positive versions of all three isoforms of SREBP. We conclude that ACS mRNA, and hence the ability of cells to activate acetate, is regulated by SREBPs in parallel with fatty acid synthesis in animal cells.
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PMID:Molecular characterization of human acetyl-CoA synthetase, an enzyme regulated by sterol regulatory element-binding proteins. 1084 99


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