Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.3.99.3 (acyl-CoA dehydrogenase)
 1,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Medium-chain acyl-CoA dehydrogenase (MCAD; acyl-CoA: (acceptor) 2,3-oxidoreductase, EC 1.3.99.3) is one of three similar enzymes that catalyze the initial step of fatty acid beta-oxidation. Definition of the primary structure of MCAD and the tissue distribution of its mRNA is of biochemical and clinical importance because of the recent recognition of inherited MCAD deficiency in humans. The MCAD mRNA nucleotide sequence was determined from two overlapping cDNA clones isolated from human liver and placental cDNA libraries, respectively. The MCAD mRNA includes a 1263-base-pair coding region and a 738-base-pair 3'-nontranslated region. A partial amino acid sequence (137 residues) determined on peptides derived from MCAD purified from porcine liver confirmed the identity of the cDNA clone. Comparison of the amino acid sequence predicted from the human MCAD cDNA with the partial protein sequence of the porcine MCAD revealed a high degree (88%) of interspecies sequence identity. RNA blot analysis shows that MCAD mRNA is expressed in a variety of rat (2.2 kilobases) and human (2.4 kilobases) tissues. Blot hybridization of RNA prepared from cultured skin fibroblasts from a patient with MCAD deficiency disclosed that mRNA was present and of similar size to MCAD mRNA derived from control fibroblasts. The isolation and characterization of MCAD cDNA is an important step in the definition of the defect underlying MCAD deficiency and in understanding its metabolic consequences.
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PMID:Nucleotide sequence of medium-chain acyl-CoA dehydrogenase mRNA and its expression in enzyme-deficient human tissue. 303 65

A procedure was developed to assay [14C]octanoate oxidation from the production of both 14CO2 and 14C-labeled acid-soluble products. Octanoic acid and its CoA and carnitine esters were eliminated from the acid-soluble products by alkaline hydrolysis of the esters and acidification and binding of the acid to Lipidex 1000. The method was evaluated with homogenates of various rat tissues and human muscles and with human fibroblasts. 14CO2 production was variable and comprised less than 3% of the total oxidation products with homogenates and 26 +/- 19% with fibroblasts. As compared to palmitate, oxidation rates of octanoate were higher in rat liver and heart homogenates, of the same magnitude in muscle homogenates, but lower in fibroblasts. The proportion of antimycin-insensitive oxidation was much lower with octanoate than with palmitate. Using the assay a case of medium-chain acyl-CoA dehydrogenase deficiency could be indicated.
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PMID:An accurate and sensitive assay of [14C]octanoate oxidation and its application on tissue homogenates and fibroblasts. 308 Oct 42

Thia- and oxaoctanoyl-CoA derivatives (substituted at the C-3 and C-4 positions) have been synthesized to prove the reductive half-reaction in the medium-chain acyl-CoA dehydrogenase from pig kidney. 3-Thiaoctanoyl-CoA binds to this flavoenzyme, forming an intense, stable, long-wavelength band (at 804 nm; extinction coefficient = 8.7 mM-1 cm-1 at pH 7.6). The intensity of this band increases about 20% from pH 6.0 to pH 8.8. This long-wavelength species probably represents a charge-transfer complex between bound acyl enolate as the donor and oxidized flavin adenine dinucleotide as the acceptor. Thus, the enzyme catalyzes alpha-proton exchange, and no long-wavelength bands are seen with 3-thiaoctyl-CoA (where the carbonyl moiety is replaced by a methylene group). 3-Oxaoctanoyl-CoA binds comparatively weakly to the dehydrogenase, with a long-wavelength band at 780 nm which is both less intense and less stable than the corresponding thia analogue. These data suggest that the enzyme can accomplish alpha-proton abstraction from certain weakly acidic acyl-CoA derivatives, without concerted transfer of a hydride equivalent to the flavin. 4-Thiaoctanoyl-CoA is dehydrogenated in the standard assay 1.5-fold faster than octanoyl-CoA. Titrations of the medium-chain dehydrogenase with the 4-thia derivative resemble those obtained with octanoyl-CoA, except for the contribution of the strongly absorbing 4-thia-trans-2-octenoyl-CoA product. The corresponding 4-oxa analogue is a much poorer substrate (10% of the rate shown by octanoyl-CoA) but again effects substantially complete reduction of the flavin chromophore in the dehydrogenase.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:The reductive half-reaction in Acyl-CoA dehydrogenase from pig kidney: studies with thiaoctanoyl-CoA and oxaoctanoyl-CoA analogues. 316 33

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

Pig kidney medium-chain acyl-CoA dehydrogenase (EC 1.3.99.3) is irreversibly and stoichiometrically inactivated by [1-14C]-2-octynoyl coenzyme A. The linkage is stable at pH 2-6, but labile under basic conditions. The inhibitor labels a unique tryptic peptide, Ile-Tyr-Gln-Ile-Tyr-Glu-Gly-Thr-Ala-Gln-Ile-Gln-Arg, close to the C-terminus of the protein. The peptide is labeled at Glu-401 with the acyl moiety of the inhibitor but does not contain detectable coenzyme A. Both the inactivation of the dehydrogenase and the appearance of an absorption band at 800 nm show large primary deuterium isotope effects using 4,4'-dideuterio-2-octynoyl-CoA (7.3 and 6.3, respectively). Thus, 2-octynoyl-CoA is a mechanism-based inactivator of the dehydrogenase and is activated by rate-limiting gamma-proton abstraction. Glutamate-401 may be the base that abstracts the pro-R alpha-proton during the dehydrogenation of normal substrates.
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PMID:2-octynoyl coenzyme A is a mechanism-based inhibitor of pig kidney medium-chain acyl coenzyme A dehydrogenase: isolation of the target peptide. 323 92

The carnitine system functions in the transport of activated acyl groups over the mitochondrial inner membrane, and is needed for oxidation of long-chain fatty acids by all mitochondria. The rate of cardiac fatty acid oxidation is determined by availability of fatty acids, oxygen and the activity of carnitine palmitoyltransferase I, which is regulated by a variety of factors. It is inhibited by malonyl-CoA, which in rat heart was found to be synthesized by acetyl-CoA carboxylase. It is also inhibited by long-chain acylcarnitine. Linoleoylcarnitine was found to be a better inhibitor than palmitoylcarnitine. The concentration of carnitine in human heart, muscle and other tissues is much higher than is needed for the optimal beta-oxidation rate. In contrast to controls, we found in several myopathic patients that extra carnitine (from 1/2 to 5 mM) caused a considerable increase in beta-oxidation rate of isolated muscle mitochondria. In some of these patients we detected medium-chain acyl-CoA dehydrogenase deficiency. Patients with primary carnitine deficiency caused by a renal carnitine leak often show cardiomyopathy, which completely disappears under carnitine therapy. Cardiomyopathy may also be the cause of secondary carnitine deficiency resulting from a mitochondrial defect in acyl-CoA metabolism, or by the mitochondrial defect itself, which may be induced by drugs or viral attack, or be the result of a genetic error. In cardiomyopathic patients with a (subclinical) myopathy, study of isolated mitochondria and homogenate from skeletal muscle may reveal a mitochondrial dysfunction, which, in some patients, is treatable by dietary measures and supplementation with vitamins, CoQ and/or carnitine. When the cause of cardiomyopathy is not known, determination of plasma carnitine and carnitine supplementation of hypocarnitinemic patients is of great therapeutic value.
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PMID:The role of the carnitine system in myocardial fatty acid oxidation: carnitine deficiency, failing mitochondria and cardiomyopathy. 331 Oct 10

Our early study of isovaleric acidemia (IVA) indicated that isovaleryl-CoA is dehydrogenated by an enzyme that is specific for isovaleryl-CoA. We subsequently identified and purified isovaleryl-CoA dehydrogenase (IVD) and 2-methyl-branched chain acyl-CoA dehydrogenase, which were previously unknown. We also purified and characterized three previously known acyl-CoA dehydrogenases. Five acyl-CoA dehydrogenases share similar molecular features and reaction mechanisms, indicating a close evolutionary relationship. Using the tritium release assay and [35S]methionine labeling/immunoprecipitation, we showed that IVA is due to a mutation of IVD. We also demonstrated that there are at least 5 distinct forms of mutant IVD, indicating an extensive molecular heterogeneity. Furthermore, we cloned cDNAs encoding IVD and medium-chain acyl-CoA dehydrogenases. The comparison of their complete primary sequences revealed a high degree of homology, indicating that these enzymes belong to a gene family, the acyl-CoA dehydrogenase family.
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PMID:Molecular basis of isovaleric acidemia and medium-chain acyl-CoA dehydrogenase deficiency. 332 38

The nature of the purple complex formed upon the addition of octanoyl-CoA to the medium chain acyl-CoA dehydrogenase from pig kidney has been addressed by chemical quenching studies. Previous work, using quenching in 0.1 M KOH, suggested that the dehydrogenation product, trans-2-octenoyl-CoA, was not a participant in reduced rat liver enzyme complexes because no octenoic acid was detected after denaturation (Y. Ikeda, D. G. Hine, K. Okamura-Ikeda and K. Tanaka (1985) J. Biol. Chem. 260, 1326-1337). However, when the octanoyl-CoA-reduced pig kidney enzyme is quenched rapidly in 2 M HCl, the ratio of trans-2-octenoyl-CoA/octanoyl-CoA released is 9/1. A milder acid denaturation procedure yields the corresponding ratio of 0.4/1, i.e., now with an excess of the saturated substrate. Similarly, quenching the pig kidney dehydrogenase in 0.1 M KOH reveals only minor levels of octenoyl chains released into the supernatant. When quenching is insufficiently rapid compared to the internal equilibration of oxidized enzyme.octanoyl-CoA and reduced enzyme.octenoyl-CoA forms, the outcome is decided by the greater kinetic lability of the oxidized enzyme species. These data are fully consistent with the original ascription that the purple species observed upon reduction of the acyl-CoA dehydrogenases with substrate represents a charge transfer complex between reduced flavin as the donor and trans-2-octenoyl-CoA as the acceptor.
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PMID:The nature of enzyme-substrate complexes in acyl-coenzyme A dehydrogenases. 335 70

This study examines the relationship between impaired fatty acid oxidation and the pathogenesis of Reye syndrome. We present a hypothesis proposing that many clinical signs of this childhood disease are caused by accumulation of unusual acyl CoA esters, precursors to deacylated metabolites found in the patients' blood and urine. A new method was developed to measure acyl CoA compounds in small human liver biopsy samples, offering several advantages over previous techniques. A major finding was an accumulation in Reye syndrome patients of short- and medium-chain acyl CoA intermediates of fatty acid and branched-chain amino acid oxidation. These metabolites included octanoyl, isovaleryl, butyryl, isobutyryl, propionyl, and methylmalonyl CoA esters. The findings were explained in a model of hepatic fatty acid oxidation involving three interrelated pathways: mitochondrial beta-oxidation, peroxisomal beta-oxidation, and omega-oxidation in the endoplasmic reticulum. The results suggest that pathogenesis in Reye syndrome stems from generalized mitochondrial damage resulting in accumulation of acyl CoA esters. High levels of these compounds lead to inhibition of mitochondrial pathways for ureogenesis, gluconeogenesis, and fatty acid oxidation. The inhibited pathways, in turn, could cause the hyperammonemia, hypoglycemia, and hypoketonemia observed in patients. The model also explains underlying biochemical differences between patients with Reye syndrome and medium-chain acyl CoA dehydrogenase deficiency, another disorder of fatty acid metabolism. Acetyl CoA levels, in the latter disease, were dramatically decreased, compared with both human controls and Reye syndrome patients.
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PMID:Relationship between unusual hepatic acyl coenzyme A profiles and the pathogenesis of Reye syndrome. 341 71

We prepared monospecific antisera in rabbits against purified rat short-, medium-, and long-chain acyl-CoA dehydrogenases, isovaleryl-CoA dehydrogenase, and ETF and tested the immunocross-reactivity to the corresponding human enzymes. Each antiserum specifically reacted with the corresponding human enzyme. When immunoprecipitates were analyzed by SDS-PAGE, the mobilities of all the human acyl-CoA dehydrogenases and ETF subunits were identical to those of the rat counterparts with a single exception. Human medium-chain acyl-CoA dehydrogenase had a mobility on SDS-PAGE slightly slower than that of rat medium-chain acyl-CoA dehydrogenase, suggesting that human medium-chain acyl-CoA dehydrogenase was 1 kDa larger than the rat counterpart. The immunocross-reactivity of the antisera, raised against the rat acyl-CoA dehydrogenases and ETF to the human counterpart, provide useful tools for the study of mutant enzymes in cells from patients with a genetic defect of acyl-CoA dehydrogenases of ETF.
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PMID:Immunoprecipitation and electrophoretic analysis of four human acyl-CoA dehydrogenases and electron transfer flavoprotein using antibodies raised against the corresponding rat enzymes. 360 93


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