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:1.3.1.8 (acyl-CoA dehydrogenase)
785 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Butyrivibrio fibrisolvens is a major butyrate-forming species in the bovine and ovine rumen. The enzymology of butyrate formation from pyruvate was investigated in cell-free extracts of B. fibrisolvens D1. Pyruvate owas oxidized to acetylcoenzyme A (CoA) in the presence of CoA.SH and benzyl viologen or flavin nucleotides. The bacterium uses thiolase, beta-hydroxybutyryl-CoA dehydrogenase, crotonase, and crotonyl-CoA reductase to form butyryl-CoA from acetyl-CoA. Reduction of acetoacetyl-CoA to beta-hydroxybutyryl-CoA was faster with NADH than with NADPH. Crotonyl-CoA was reduced to butyryl-CoA by NADH, but not by NADPH, only in the presence of flavin nucleotides. Reduction of flavin nucleotides by NADH was much slower than the flavin-dependent reduction of crotonyl-CoA. This indicates that flavoproteins rather than free flavin participated in the reduction of crotonyl-CoA. Butyryl-CoA was converted to butyrate by phosphate butyryl transferase and butyrate kinase.
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PMID:Enzymology of butyrate formation by Butyrivibrio fibrisolvens. 3 24

Freeze-thawed rat liver mitochondria were extensively washed with potassium phosphate, pH 7.5, and the residue was extracted with 10 mM potassium phosphate, pH 7.5, 1% (w/v) sodium cholate, 0.5 M KCl. The four beta-oxidation enzyme activities of the washes and the last extract were assayed with substrates of various carbon chain lengths. Our data suggest that the last extract contains a novel acyl-CoA dehydrogenase and long-chain 3-hydroxyacyl-CoA dehydrogenase. A novel acyl-CoA dehydrogenase was purified. The molecular masses of the native enzyme and the subunit were estimated to be 150 and 71 kDa, respectively. One mole of enzyme contained 2 mole of FAD. These properties and immunochemical properties of the enzyme differed from those of three other acyl-CoA dehydrogenases: short-, medium-, and long-chain acyl-CoA dehydrogenases. Carbon chain length specificity of the enzyme differed from that of other acyl-CoA dehydrogenases. The enzyme was active toward CoA esters of long- and very-long-chain fatty acids, but not toward those of medium- and short-chain fatty acids. The specific enzyme activity was greater than 10 times that of long-chain acyl-CoA dehydrogenase when palmitoyl-CoA was used as substrate. We propose the name "very-long-chain acyl-CoA dehydrogenase" for this enzyme.
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PMID:Novel fatty acid beta-oxidation enzymes in rat liver mitochondria. I. Purification and properties of very-long-chain acyl-coenzyme A dehydrogenase. 173 Jun 32

The free two-electron-reduced form of medium-chain acyl-CoA dehydrogenase is reoxidized by 120 microM molecular oxygen (50 mM phosphate buffer, pH 7.6, 2 degrees C) with a half-time of approximately 7 s. Reoxidation yields hydrogen peroxide as a major product with only traces of the superoxide anion. In contrast, enzyme reduced with octanoyl-CoA is extremely slowly reoxidized oxygen, and so a series of 14 different substrate analogues have been tested to assess the structural factors responsible for this effect. Complexes with redox-inactive ligands such as 3-thia- and 2-azaoctanoyl-CoA lead to an approximately 3000-fold slowing of the rate of reoxidation of the free dihydroflavin form of the enzyme. Comparable ligands lacking the thioester carbonyl function are much less effective with rates some 1.3-4-fold slower than the free enzyme. The strong suppression of oxygen reactivity observed with certain ligands is probably not simply a steric effect but may reflect desolvation of the active site and consequent destabilization of the superoxide anion intermediate formed during reoxidation of the flavin. The profound differences in oxygen reactivity between acyl-CoA dehydrogenase and acyl-CoA oxidase and the unusual stability of certain flavoprotein semiquinones in air are discussed in terms of these thermodynamic and kinetic arguments.
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PMID:Reactivity of medium-chain acyl-CoA dehydrogenase toward molecular oxygen. 186 64

Pig kidney general acyl-CoA dehydrogenase (GAD) can be reduced by butyryl-CoA to form reduced enzyme and crotonyl-CoA. This reaction is reversible. Stopped-flow, kinetic investigations on GAD have been made, using the following reaction pairs: oxidized GAD/butyryl-CoA, oxidized GAD/crotonyl-CoA, oxidized GAD/alpha,beta-dideuteriobutyryl-CoA, reduced GAD/butyryl-CoA, and reduced GAD/crotonyl-CoA (in 50 mM potassium phosphate buffer, pH 7.6 at 4 degrees C). Reduction of GAD by butyryl-CoA is triphasic. The slowest phase is 100-fold slower than the preceding phase and appears to represent a secondary process not directly related to the primary reduction events. The first two fast phases are responsible for reduction of GAD. Reduction proceeds via a reduced enzyme/crotonyl-CoA charge-transfer complex. alpha, beta-Dideuteriobutyryl-CoA elicits a major deuterium isotope effect (15-fold) on the reduction reaction. Oxidation of GAD by crotonyl-CoA is biphasic. Oxidation proceeds via the same reduced enzyme/crotonyl-CoA charge-transfer complex seen during reduction. The oxidation reaction ends in a mixture composed largely of oxidized GAD species. From the data, we constructed a mechanism for the reduction/oxidation of GAD by butyryl-CoA/crotonyl-CoA. This mechanism was then used to simulate all of the observed kinetic time course data, using a single set of kinetic parameters. A close correspondence between the observed and simulated data was obtained.
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PMID:Oxidation-reduction of general acyl-CoA dehydrogenase by the butyryl-CoA/crotonyl-CoA couple. A new investigation of the rapid reaction kinetics. 321 56

The flavoenzyme pig kidney general acyl-CoA dehydrogenase (EC 1.3.99.3) is inactivated by cyclohexane-1,2-dione in borate buffer in a reaction that exhibits pseudo-first-order kinetics. Strong protection is afforded by the substrate octanoyl-CoA, as well as by heptadecyl-CoA, a potent competitive inhibitor of the dehydrogenase that does not reduce enzyme flavin. Enzyme exhibiting 10% residual activity in borate buffer contains about 1.3 modified arginine residues per flavin molecule. Very little reduction of the modified enzyme in borate buffer occurs at high concentrations of octanoyl-CoA, in marked contrast with the stoicheiometric reduction of the native enzyme. However, in phosphate buffer alone, the modified enzyme exhibits 55% residual activity and, although binding of substrate is still seriously impaired (apparent Kd=14 microM), excess substrate effects the formation of the characteristic reduced flavin X enoyl-CoA charge-transfer complex. These results suggest that the susceptible arginine residue, though not catalytically essential, is probably within the acyl-CoA-binding site of general acyl-CoA dehydrogenase.
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PMID:Modification of an arginine residue in pig kidney general acyl-coenzyme A dehydrogenase by cyclohexane-1,2-dione. 716 2

Aspects of the binding and dehydrogenation of acyl-CoA thiol esters by the general acyl-CoA dehydrogenase from pig liver were investigated using a dead-end inhibitor, S-octyl-CoA, several alternate substrates, and three active site-directed inhibitors. Experiments with S-octyl-CoA indicate that the carbonyl group of acyl-CoA thiol esters is not absolutely required for binding to the enzyme. However, the mode of binding of the 8-carbon thiol ether can be distinguished from the mode of binding of the enoyl-CoA product, octenoyl-CoA. Octanoyl pantetheine, octanoyl-etheno-CoA, and octanoyl-3'-dephospho-CoA are alternate substrates of the dehydrogenase. Steady state kinetic constants obtained with these alternate substrates indicate that the adenosine 5'-diphosphate, but not the 3'-phosphate, of the nucleotide moiety of acyl-CoA substrates contribute to the tight binding of the substrates. The substrate analogs 3'-butynoyl-CoA and 3-octynoyl-CoA are active site-directed, mechanism-based irreversible inhibitors of the dehydrogenase. These inhibitors covalently modify the apoprotein rather than the flavin. This finding and the fact that 2,3-octadienoyl-CoA also completely and irreversibly inhibits the enzyme indicate that th 3-acetylenic thiol esters inhibit the enzyme by a mechanism involving: (1) base-catalyzed abstraction of a protein at C-2 followed by isomerization to the allene carbanion, (2) protonation of the carbanion, and (3) attack of a nucleophile in the enzyme-active site on C-3 of the 2,3-dienoyl-CoA. The data show that the alkynoyl-CoA's are activated and bound at the active site of the enzyme. The results suggest that abstraction of a proton at C-2 of acyl-CoA substrates is the initial step in the catalytic pathway of dehydrogenation of substrates by the enzyme.
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PMID:Enzyme-activated inhibitors, alternate substrates, and a dead end inhibitor of the general acyl-CoA dehydrogenase. 744 May 36

We undertook a comparative investigation of the medium-chain fatty acyl-CoA dehydrogenase (MCAD)-catalyzed reaction utilizing indole-, furyl-, and 4-(dimethylamino)phenyl-substituted propionyl- and acryloyl-CoAs as potential substrate/product pairs. All these propionyl-CoA derivatives undergo MCAD-catalyzed conversion into their corresponding acryloyl-CoAs via both "dehydrogenase" (in the presence of "organic" electron acceptors) and "oxidase" (buffer-dissolved oxygen serving as the electron acceptor) pathways [Johnson, J. K., Wang, Z. X., & Srivastava, D. K. (1992) Biochemistry 31, 10564-10575]. The steady-state kinetic parameters for the enzyme utilizing these substrates reveal that the KmS (for the CoA substrates) and kcatS for the dehydrogenase reaction are at least an order of magnitude higher than those for the oxidase reaction. As with the CoA substrates, the enzyme catalyzes the conversion of indolepropionyl pantetheine phosphate (IPPP) into indoleacryloyl pantetheine phosphate (IAPP) via these two pathways. However, with IPPP as substrate, the Km (for IPPP) and kcat values of the dehydrogenase and oxidase reactions are the same. These, coupled with the spectral changes of the enzyme-product complexes as well as the binding affinities of the enzyme-substrate/product complexes, lead to the following conclusions: (1) The aromatic/heterocyclic group-containing substrates are converted into their corresponding products via both the dehydrogenase and the oxidase pathways. (2) The 3',5'-ADP moiety of the CoA thioester provides a significant fraction of the total binding energy in stabilizing the enzyme-substrate/product complexes.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:"Dehydrogenase" and "oxidase" reactions of medium-chain fatty acyl-CoA dehydrogenase utilizing chromogenic substrates: role of the 3',5'-adenosine diphosphate moiety of the coenzyme A thioester in catalysis. 771 65

A high-performance liquid chromatographic method is presented for the determination of urinary acylcarnitines. After solid phase extraction on silica columns the acylcarnitines are converted to 4'-bromophenacyl esters with 4'-bromophenacylbromide in the presence of N,N-diisopropylethylamine. Complete derivatization was achieved at 37 degrees C within 30 min. The 4'-bromophenacyl esters were separated by high-performance liquid chromatography on a Hypersil BDS C8 reversed-phase column with a binary gradient containing varying proportions of acetonitrile, water and 0.1 M triethylamine phosphate buffer. Essentially baseline separation was obtained with a standard mixture containing 4'-bromophenacyl esters of carnitine and synthetic acylcarnitines of increasing chain length ranging from acetyl- to palmitoylcarnitine. The method was used to obtain urinary acylcarnitine profiles from patients with propionic, methylmalonic and isovaleric acidemia and with medium-chain and multiple acyl-CoA dehydrogenase deficiency. Quantification of the acylcarnitines was achieved using undecanoylcarnitine as internal standard.
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PMID:Determination of acylcarnitines in urine of patients with inborn errors of metabolism using high-performance liquid chromatography after derivatization with 4'-bromophenacylbromide. 822 73

We investigated the binding of octenoyl-CoA to pig kidney medium chain acyl-CoA dehydrogenase (MCAD) by isothermal titration microcalorimetry under a variety of experimental conditions. At 25 degrees C in 50 mM phosphate buffer at pH 7.6 (ionic strength of 175 mM), the binding is characterized by the stoichiometry (n) of 0.89 mole of octenoyl-CoA/(mole of MCAD subunit), delta G = -8.75 kcal/mol, delta H = -10.3 kcal/mol, and delta S = -5.3 cal mol(-1) K(-1), suggesting that formation of MCAD-octenoyl-CoA is enthalpically driven. By employing buffers with various ionization enthalpies, we discerned that formation of the MCAD-octenoyl-CoA complex, at pH 7.6, accompanies abstraction (consumption) of 0.52 +/- 0.15 proton/(MCAD subunit) from the buffer media. We studied the effects of pH, ionic strength, and temperature on the thermodynamics of MCAD-octenoyl-CoA interaction. Whereas the ionic strength does not significantly influence the above interaction, the pH of the buffer media exhibits a pronounced effect. The pH dependence of the association constant of MCAD +octenoyl-CoA <==> MCAD-octenoyl-CoA yields a pKa for the free enzyme of 6.2. Among thermodynamic parameters, whereas delta G remains invariant as a function of temperature, delta H and deltaS(standard) both decrease with an increase in temperature. At temperatures of < 25 degrees C, delta G is dominated by favorable entropic contributions. As the temperature increases, the entropic contributions progressively decrease, attain a value of zero at 23.8 degrees C, and then becomes unfavorable. During this transition, the enthalpic contributions become progressively favorable, resulting in an enthalpy-entropy compensation. The temperature dependence of delta H yields the heat capacity change (delta Cp(0)) of -0.37 +/- 0.05 kcal mol(-1) K(-1), attesting to the fact that the binding of octenoyl-CoA to MCAD is primarily dominated by the hydrophobic forces. The thermodynamic data presented herein are rationalized in light of structural-functional relationships in MCAD catalysis.
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PMID:Isothermal titration microcalorimetric studies for the binding of octenoyl-CoA to medium chain acyl-CoA dehydrogenase. 917 51

Mature medium chain acyl-CoA dehydrogenase isolated from pig kidney (pkMCADH) and originating from mitochondria carries a phosphate group as demonstrated by 31P-NMR-spectroscopy and chemical analysis. Two broad resonances at -6.3 and -8 ppm are observed and are assigned to the pyrophosphate group of the cofactor FAD. A third, narrow resonance at 4.65 ppm indicates the presence of a phosphomonoester residue. Chemical analysis of intact pkMCADH shows the presence of 3 +/- 0.3 phosphates, those of FAD and of an additional covalently attached phosphate. With recombinant, human wild type MCADH expressed in and purified from E. coli only the two FAD phosphates (2 +/- 0.35) are found. Similarly, pkMCADH which has been converted to the apoenzyme and reconstituted to holoenzyme also contains 2 +/- 0.4 phosphates. The covalently bound phosphate can be hydrolyzed by phosphatase and subsequently removed by dialysis. The phosphate group has no detectable effect on the catalytic activity of the MCADH measured with artificial and natural electron acceptors such as pig electron transferring flavoprotein. However, phosphorylation has a marked effect on protein solubility which is +5-fold lower for the dephosphorylated protein.
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PMID:Medium-chain acyl CoA dehydrogenase: evidence for phosphorylation. 942 98


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