Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:6.2.1.1 (ACS)
78,556 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The bifunctional CO dehydrogenase/acetyl-CoA synthase (CODH/ACS) plays a central role in the Wood-Ljungdahl pathway of autotrophic CO(2) fixation. A recent structure of the Moorella thermoacetica enzyme revealed that the ACS active site contains a [4Fe-4S] cluster bridged to a binuclear Cu-Ni site. Here, biochemical and x-ray absorption spectroscopic (XAS) evidence is presented that the copper ion at the M. thermoacetica ACS active site is essential. Depletion of copper correlates with reduction in ACS activity and in intensity of the "NiFeC" EPR signal without affecting either the activity or the EPR spectroscopic properties associated with CODH. In contrast, Zn content is negatively correlated with ACS activity without any apparent relationship to CODH activity. Cu is also found in the methanogenic CODH/ACS from Methanosarcina thermophila. XAS studies are consistent with a distorted Cu(I)-S(3) site in the fully active enzyme in solution. Cu extended x-ray absorption fine structure analysis indicates an average Cu-S bond length of 2.25 A and a metal neighbor at 2.65 A, consistent with the Cu-Ni distance observed in the crystal structure. XAS experiments in the presence of seleno-CoA reveal a Cu-S(3)Se environment with a 2.4-A Se-Cu bond, strongly implicating a Cu-SCoA intermediate in the mechanism of acetyl-CoA synthesis. These results indicate an essential and functional role for copper in the CODH/ACS from acetogenic and methanogenic organisms.
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PMID:Functional copper at the acetyl-CoA synthase active site. 1258 21

Acetyl-coenzyme A synthetase catalyzes the two-step synthesis of acetyl-CoA from acetate, ATP, and CoA and belongs to a family of adenylate-forming enzymes that generate an acyl-AMP intermediate. This family includes other acyl- and aryl-CoA synthetases, firefly luciferase, and the adenylation domains of the modular nonribosomal peptide synthetases. We have determined the X-ray crystal structure of acetyl-CoA synthetase complexed with adenosine-5'-propylphosphate and CoA. The structure identifies the CoA binding pocket as well as a new conformation for members of this enzyme family in which the approximately 110-residue C-terminal domain exhibits a large rotation compared to structures of peptide synthetase adenylation domains. This domain movement presents a new set of residues to the active site and removes a conserved lysine residue that was previously shown to be important for catalysis of the adenylation half-reaction. Comparison of our structure with kinetic and structural data of closely related enzymes suggests that the members of the adenylate-forming family of enzymes may adopt two different orientations to catalyze the two half-reactions. Additionally, we provide a structural explanation for the recently shown control of enzyme activity by acetylation of an active site lysine.
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PMID:The 1.75 A crystal structure of acetyl-CoA synthetase bound to adenosine-5'-propylphosphate and coenzyme A. 1262 52

The influence of residual ethanol on metabolism of food grade Gluconacetobacter xylinus I 2281 was investigated during controlled cultivations on 35 g/l glucose and 5 g/l ethanol. Bacterial growth was strongly reduced in the presence of ethanol, which is unusual for acetic acid bacteria. Biomass accumulated only after complete oxidation of ethanol to acetate and carbon dioxide. In contrast, bacterial growth initiated without delay on 35 g/l glucose and 5 g/l acetate. It was found that acetyl CoA was activated by the acetyl coenzyme A synthetase (Acs) pathway in parallel with the phosphotransacetylase (Pta)-acetate kinase (Ack) pathway. The presence of ethanol in the culture medium strongly reduced Pta activity while Acs and Ack remained active. A carbon balance calculation showed that the overall catabolism could be divided into two independent parts: upper glycolysis linked to glucose catabolism and lower glycolysis liked to ethanol catabolism. This calculation showed that the carbon flux through the tricarboxylic cycle is lower on ethanol than on acetate. This corroborated the diminution of carbon flux through the Pta-Ack pathway due to the inhibition of Pta activity on ethanol.
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PMID:Influence of residual ethanol concentration on the growth of Gluconacetobacter xylinus I 2281. 1269 73

The structure of carbon monoxide dehydrogenase/acetyl-coenzyme A synthase (CODH/ACS), a central enzyme in the anaerobic metabolism of acetyl-coenzyme A (acetyl-CoA), has been solved to a resolution of 2.2A. The active-site metal cluster responsible for catalyzing acetyl C-C bond synthesis and cleavage, designated the A center, was identified as an Fe(4)S(4) iron sulfur cluster with one of its cysteine thiolates acting as a bridge to an adjacent binuclear metal site. Nickel was found at one position in the binuclear site and the other metal was indicated to be copper - a surprising result, implying a previously unrecognized role for copper. Details of the A center provided new insight into the unusual organometallic mechanism of acetyl C-C bond formation and cleavage, with substantial conformational changes indicated for binding of the large methylcorrinoid protein substrate, and a unique intramolecular channel acting to contain carbon monoxide within the protein and transfer it to the site needed for acetyl-CoA synthesis.
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PMID:Acetate C-C bond formation and decomposition in the anaerobic world: the structure of a central enzyme and its key active-site metal cluster. 1276 30

The aim of this work was to understand the steps controlling the process of biotransformation of trimethylamonium compounds into L(-)-carnitine by Escherichia coli and the link between the central carbon or primary and the secondary metabolism expressed. Thus, the enzyme activities involved in the biotransformation process of crotonobetaine into L(-)-carnitine (crotonobetaine hydration reaction and crotonobetaine reduction reaction), in the synthesis of acetyl-CoA (pyruvate dehydrogenase, acetyl-CoA synthetase, and ATP:acetate phosphotransferase) and in the distribution of metabolites for the tricarboxylic acid (isocitrate dehydrogenase) and glyoxylate (isocitrate lyase) cycles, were followed in batch with both growing and resting cells and during continuous cell growth in stirred-tank and high-cell-density membrane reactors. In addition, the levels of carnitine, crotonobetaine, gamma-butyrobetaine, ATP, NADH/NAD(+), and acetyl-CoA/CoA ratios were measured to determine how metabolic fluxes were distributed in the catabolic system. The results provide the first experimental evidence demonstrating the important role of the glyoxylate shunt during biotransformation of resting cells and the need for high levels of ATP to maintain metabolite transport and biotransformation (2.1 to 16.0 mmol L cellular/mmol ATP L reactor h). Moreover, the results obtained for the pool of acetyl-CoA/CoA indicate that it also correlated with the biotransformation process. The main metabolic pathway operating during cell growth in the high cell-density membrane reactor was that related to isocitrate dehydrogenase (during start-up) and isocitrate lyase (during steady-state operation), together with phosphotransacetylase and acetyl-CoA synthetase. More importantly, the link between central carbon and L(-)-carnitine metabolism at the level of the ATP pool was also confirmed.
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PMID:Link between primary and secondary metabolism in the biotransformation of trimethylammonium compounds by escherichia coli. 1459 81

Acetyl-CoA synthase (ACS identical with ACS/CODH identical with CODH/ACS) from Moorella thermoacetica catalyzes the synthesis of acetyl-CoA from CO, CoA, and a methyl group of a corrinoid-iron-sulfur protein (CoFeSP). A time lag prior to the onset of acetyl-CoA production, varying from 4 to 20 min, was observed in assay solutions lacking the low-potential electron-transfer agent methyl viologen (MV). No lag was observed when MV was included in the assay. The length of the lag depended on the concentrations of CO and ACS, with shorter lags found for higher [ACS] and sub-saturating [CO]. Lag length also depended on CoFeSP. Rate profiles of acetyl-CoA synthesis, including the lag phase, were numerically simulated assuming an autocatalytic mechanism. A similar reaction profile was monitored by UV-vis spectrophotometry, allowing the redox status of the CoFeSP to be evaluated during this process. At early stages in the lag phase, Co(2+)FeSP reduced to Co(+)FeSP, and this was rapidly methylated to afford CH(3)-Co(3+)FeSP. During steady-state synthesis of acetyl-CoA, CoFeSP was predominately in the CH(3)-Co(3+)FeSP state. As the synthesis rate declined and eventually ceased, the Co(+)FeSP state predominated. Three activation reductive reactions may be involved, including reduction of the A- and C-clusters within ACS and the reduction of the cobamide of CoFeSP. The B-, C-, and D-clusters in the beta subunit appear to be electronically isolated from the A-cluster in the connected alpha subunit, consistent with the ~70 A distance separating these clusters, suggesting the need for an in vivo reductant that activates ACS and/or CoFeSP.
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PMID:Autocatalytic activation of acetyl-CoA synthase. 1501 40

4-Chlorobenzoate:CoA ligase (CBAL) is a member of a family of adenylate-forming enzymes that catalyze two-step adenylation and thioester-forming reactions. In previous studies, we have provided structural evidence that members of this enzyme family (exemplified by acetyl-CoA synthetase) use a large domain rotation to catalyze the respective partial reactions [A. M. Gulick, V. J. Starai, A. R. Horswill, K. M. Homick, and J. C. Escalante-Semerena, (2003) Biochemistry 42, 2866-2873]. CBAL catalyzes the synthesis of 4-chlorobenzoyl-CoA, the first step in the 4-chlorobenzoate degredation pathway in PCB-degrading bacteria. We have solved the 2.0 A crystal structure of the CBAL enzyme from Alcaligenes sp. AL3007 using multiwavelength anomalous dispersion. The results demonstrate that in the absence of any ligands, or bound to the aryl substrate 4-chlorobenzoate, the enzyme adopts the conformation poised for catalysis of the adenylate-forming half-reaction. We hypothesize that coenzyme A binding is required for stabilization of the alternate conformation, which catalyzes the 4-CBA-CoA thioester-forming reaction. We have also determined the structure of the enzyme bound to the aryl substrate 4-chlorobenzoate. The aryl binding pocket is composed of Phe184, His207, Val208, Val209, Phe249, Ala280, Ile303, Gly305, Met310, and Asn311. The structure of the 4-chlorobenzoate binding site is discussed in the context of the binding sites of other family members to gain insight into substrate specificity and evolution of new function.
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PMID:Crystal structure of 4-chlorobenzoate:CoA ligase/synthetase in the unliganded and aryl substrate-bound states. 1523 75

ADP-forming acetyl-CoA synthetase (ACD), the novel enzyme of acetate formation and energy conservation in archaea Acety - CoA + ADP + Pi<==>acetate + ATP CoA), has been studied only in few hyperthermophilic euryarchaea. Here, we report the characterization of two ACDs with unique molecular and catalytic features, from the mesophilic euryarchaeon Haloarcula marismortui and from the hyperthermophilic crenarchaeon Pyrobaculum aerophilum. ACD from H. marismortui was purified and characterized as a salt-dependent, mesophilic ACD of homodimeric structure (166 kDa). The encoding gene was identified in the partially sequenced genome of H. marismortui and functionally expressed in Escherichia coli. The recombinant enzyme was reactivated from inclusion bodies following solubilization and refolding in the presence of salts. The ACD catalyzed the reversible ADP- and Pi-dependent conversion of acetyl-CoA to acetate. In addition to acetate, propionate, butyrate, and branched-chain acids (isobutyrate, isovalerate) were accepted as substrates, rather than the aromatic acids, phenylacetate and indol-3-acetate. In the genome of P. aerophilum, the ORFs PAE3250 and PAE 3249, which code for alpha and beta subunits of an ACD, overlap each other by 1 bp, indicating a novel gene organization among identified ACDs. The two ORFs were separately expressed in E. coli and the recombinant subunits alpha (50 kDa) and beta (28 kDa) were in-vitro reconstituted to an active heterooligomeric protein of high thermostability. The first crenarchaeal ACD showed the broadest substrate spectrum of all known ACDs, catalyzing the conversion of acetyl-CoA, isobutyryl-CoA, and phenylacetyl-CoA at high rates. In contrast, the conversion of phenylacetyl-CoA in euryarchaeota is catalyzed by specific ACD isoenzymes.
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PMID:Unusual ADP-forming acetyl-coenzyme A synthetases from the mesophilic halophilic euryarchaeon Haloarcula marismortui and from the hyperthermophilic crenarchaeon Pyrobaculum aerophilum. 1534 Jul 86

Carbon monoxide dehydrogenase/acetyl-CoA synthase (CODH/ACS) is a bifunctional enzyme which enables archaea and bacteria to grow autotrophically on CO and hydrogen/carbon dioxide using the Wood-Ljundahl pathway. CO produced from reduction of carbon dioxide by CODH is transferred to the active site of ACS through an intramolecular tunnel, where it combines with Coenzyme A and a methyl cation to produce acetyl-CoA. The active site of ACS contains a single [4Fe-4S] cluster bridged by a cysteine sulfur atom to a binuclear center. The binuclear center is composed of two Ni atoms bridged by two separate cysteine sulfurs. The Ni site attached to the [4Fe-4S] is referred to as proximal Ni, while the other Ni atom, which assumes a square-planar geometry, is referred to as the distal site. We report the characterization of the carbonylated form of highly active (0.67 spins/mol) heterologously expressed monomeric ACS from C. hydrogenoformans in E. coli by rapid-freeze quench EPR (RFQ-EPR) and stopped-flow infrared (SF-IR) spectroscopies. The reaction of ACS with CO produces a single metal-carbonyl species whose formation rate, measured by SF-IR, correlates with the rate of formation, measured by RFQ-EPR, of the paramagnetic state of the enzyme (NiFeC species). These results indicate that the NiFeC species is the predominant form observed in solution when ACS reacts with CO. The NiFeC species contains the proximal Ni in the +1 redox state and the [4Fe-4S] cluster in the 2+ state, thus there is no evidence for either a Ni(0) or a Ni(II) state in the active carbonylated form of the enzyme.
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PMID:EPR and infrared spectroscopic evidence that a kinetically competent paramagnetic intermediate is formed when acetyl-coenzyme A synthase reacts with CO. 1619 Jul 5

We established a novel enzyme-catalyzed poly(3-hydroxybutyrate) [P(3HB)] synthesis system capable of recycling CoA on the basis of the P(3HB) biosynthetic pathway in Ralstonia eutropha. The system includes purified beta-ketothiolase (PhaA), NADPH-dependent acetoacetyl-CoA reductase (PhaB), PHA synthase (PhaC), acetyl-CoA synthetase (Acs) and glucose dehydrogenase (GDH). In this system, acetyl-CoA was synthesized from acetate and CoA by Acs and ATP, and then two molecules of acetyl-CoA were condensed by PhaA to synthesize acetoacetyl-CoA, which was converted to (R)-3-hydroxybutyryl-CoA (3HBCoA) by PhaB and NADPH. The 3HBCoA was polymerized by PhaC and converted to P(3HB). In this system, the CoA molecules that were released during the condensation and polymerization reactions catalyzed by PhaA and PhaC, respectively, were reused successfully for the synthesis of acetyl-CoA. In addition, NADPH, which was consumed in the reduction of acetoacetyl-CoA, was regenerated by the action of GDH. In this system, the yield of P(3HB) synthesized from acetate as the substrate was 5.6 mg in a 5-ml reaction mixture, and the weight-average molecular weight and polydispersity were 6.64 x 10(6) and 1.36, respectively. Furthermore, CoA was reused at least 26 times, and NADPH was also regenerated at least 26 times during 24 h of reaction.
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PMID:Enzyme-catalyzed poly(3-hydroxybutyrate) synthesis from acetate with CoA recycling and NADPH regeneration in Vitro. 1623 16


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