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M Pourfarzam

Publications and source records attributed to M Pourfarzam.

At least 37 records · Page 2Linked to original sources
PubMed1996

Mammalian mitochondrial beta-oxidation.

The enzymic stages of mammalian mitochondrial beta-oxidation were elucidated some 30-40 years ago. However, the discovery of a membrane-associated multifunctional enzyme of beta-oxidation, a membrane-associated acyl-CoA dehydrogenase and characterization of the carnitine palmitoyl transferase system at the protein and at the genetic level has demonstrated that the enzymes of the system itself are incompletely understood. Deficiencies of many of the enzymes have been recognized as important causes of disease. In addition, the study of these disorders has led to a greater understanding of the molecular mechanism of beta-oxidation and the import, processing and assembly of the beta-oxidation enzymes within the mitochondrion. The tissue-specific regulation, intramitochondrial control and supramolecular organization of the pathway is becoming better understood as sensitive analytical and molecular techniques are applied. This review aims to cover enzymological and organizational aspects of mitochondrial beta-oxidation together with the biochemical aspects of inherited disorders of beta-oxidation and the intrinsic control of beta-oxidation.

Acyl-CoA Dehydrogenases↗
PubMed1996

The effect of respiratory chain impairment of beta-oxidation in rat heart mitochondria.

Cardiac ischaemia leads to an inhibition of beta-oxidation flux and an accumulation of acyl-CoA and acyl-carnitine esters in the myocardium. However, there remains some uncertainty as to which esters accumulate during cardiac ischaemia and therefore the site of inhibition of beta-oxidation [Moore, Radloff, Hull and Sweely (1980) Am. J. Physiol. 239, H257-H265; Latipää (1989) J. Mol. Cell. Cardiol. 21, 765-771]. When beta-oxidation of hexadecanoyl-CoA in state III rat heart mitochondria was inhibited by titration of complex III activity, flux measured as 14CO2 release, acid-soluble radioactivity or as acetyl-carnitine was progressively decreased. Low concentrations of myxothiazol caused reduction of the ubiquinone pool whereas the NAD+/NADH redox state was less responsive. Measurement of the CoA and carnitine esters generated under these conditions showed that there was a progressive decrease in the amounts of chain-shortened saturated acyl esters with increasing amounts of myxothiazol. The concentrations of 3-hydroxyacyl and 2-enoyl esters, however, were increased between 0 and 0.2 microM myxothiazol but were lowered at higher myxothiazol concentrations. More hexadecanoyl-CoA and hexadecanoyl-carnitine were present with increasing concentrations of myxothiazol. We conclude that 3-hydroxyacyl-CoA dehydrogenase and acyl-CoA dehydrogenase activities are inhibited by reduction of the ubiquinone pool, and that this explains the confusion over which esters of CoA and carnitine accumulate during cardiac ischaemia. Furthermore these studies demonstrate that the site of the control exerted by the respiratory chain over beta-oxidation is shifted depending on the extent of the inhibition of the respiratory chain.

Acetylcarnitine↗
PubMed1995

Effects of added l-carnitine, acetyl-CoA and CoA on peroxisomal beta-oxidation of [U-14C]hexadecanoate by isolated peroxisomal fractions.

(1) During peroxisomal beta-oxidation of [U-14C]hexadecanoate, at concentrations higher than 100 microM, long-chain 3-oxoacyl-CoA-esters and 3-oxobutyryl-CoA accumulate. Only 3-oxobutyryl-CoA accumulates at a low concentration of [U-14C]hexadecanoate. Accumulation of long chain 3-oxoacyl-CoA esters is most extensive when the supply of CoA can be considered limiting for beta-oxidation. (2) Added acetyl-CoA was found to inhibit peroxisomal beta-oxidation. This inhibition was not significantly relieved by added L-carnitine and carnitine acetyltransferase (EC 2.3.17). (3) Added L-carnitine, at concentrations below 0.2 mM, was found to stimulate peroxisomal beta-oxidation of [U-14C]hexadecanoate by up to 20%, causing the conversion of acetyl-CoA into acetylcarnitine. Higher concentrations of L-carnitine were progressively inhibitory to beta-oxidation. This effect was specific for L-carnitine as both D-carnitine and aminocarnitine neither caused stimulation at low concentrations, nor inhibition at higher concentrations. Added L-carnitine caused accumulation of acylcarnitines of chain-lengths ranging from 4 to 16 carbon-atoms. The inhibition observed with higher concentrations of added L-carnitine is likely due to conversion of [U-14C]hexadecanoate into [U-14C]hexadecanoylcarnitine. (4) Low concentrations of added hexadecanoylcarnitine was shown to inhibit peroxisomal beta-oxidation by about 15%, while added acetylcarnitine did not inhibit at concentrations up to 100 microM. (5) These data are interpreted to indicate significant control being exerted on flux at the stage of thiolysis either directly by means of CoA availability, or indirectly by means of the rate of acetyl-CoA generation.

Acetyl Coenzyme A↗
PubMed1995

Impaired ketogenesis in fructose-1,6-bisphosphatase deficiency: a pitfall in the investigation of hypoglycaemia.

Intermediary metabolite concentrations were measured in blood during fasting in two patients with fructose-1,6-bisphosphatase deficiency. Hypoglycaemia was accompanied by markedly raised levels of plasma free fatty acids, without the expected degree of ketosis. This suggests that there is secondary impairment of ketogenesis in this condition, and could lead to diagnostic confusion.

3-Hydroxybutyric Acid↗
PubMed1995

Neonatal hyperammonemia caused by a defect of carnitine-acylcarnitine translocase.

Carnitine-acylcarnitine translocase deficiency is a newly recognized inborn error of metabolism that involves transport of long-chain fatty acids into mitochondria, which in turn impairs mitochondrial beta-oxidation, and ketogenesis. We report a new familial example; the affected twins had neonatal distress, hyperammonemia, and transient intracardiac conduction defects. Clinical and biochemical analysis of both our patients and the two previously reported patients revealed that this inherited defect could be manifested during the neonatal period without any of the signs classically associated with fatty oxidation defects. In contrast, all four patients had sustained and "isolated" hyperammonemia, which could be misinterpreted as being caused by urea cycle defects. We conclude that carnitine-acylcarnitine translocase deficiency is a potential differential diagnosis in neonates with unexplained neonatal hyperammonemia. Cardiac and muscle involvement may represent further early pivotal symptoms.

Ammonia↗
PubMed1995

Fatty acid oxidation in peripheral blood cells: characterization and use for the diagnosis of defects of fatty acid oxidation.

Disorders of mitochondrial fatty acid oxidation are increasingly recognized as an important group of inborn errors of metabolism that are associated with a significant, but easily preventable, morbidity and mortality in children. However, diagnosis is often delayed because there is no easily applied method that detects all defects. Therefore, we have characterized the acylcarnitine intermediates of fatty acid oxidation in peripheral blood cells from healthy control volunteers and patients with four different defects. After selective permeabilization with histone II AS, a novel permeabilizing agent, the cells were incubated with [U-14C]hexadecanoate and beta-oxidation flux and the acylcarnitine esters formed were measured. Blood cells from the control population produced large amounts of 3-hydroxyacylcarnitines and 2-enoylcarnitine esters, in addition to saturated acylcarnitine esters. This result is different from that found in other tissues (fibroblasts and muscle), where only saturated acylcarnitine esters could be detected. In blood cells from patients with defects of enzymes involved in long-chain fatty acid oxidation, flux was significantly reduced at 15 to 20% of control values (7.1 +/- 2.3 nmol C2 units formed per minute per International Unit of citrate synthase activity). There was a characteristic accumulation of acylcarnitines that was pathognomonic for the site of the defect. Thus, analysis of beta-oxidation intermediates from blood cells allows unequivocal identification of the four most common beta-oxidation defects.

Acylation↗
PubMed1994

Very long-chain acyl coenzyme A dehydrogenase deficiency presenting with exercise-induced myoglobinuria.

A young man presented with recurrent episodes of muscle pain and myoglobinuria after prolonged exercise or fasting. Studies on isolated muscle mitochondria showed slow flux through beta-oxidation and the presence of only saturated long-chain acyl coenzyme A (acyl-CoA) esters. These results strongly suggested a defect in the dehydrogenation of long-chain acyl-CoA esters that we confirmed by measurement of enzyme activity in muscle and platelet mitochondrial fractions and fibroblast homogenates. In all tissues studied from the patient, the enzyme activity was approximately 10% of control values with acyl-CoA esters from C16-C22 as substrates. We investigated the intramitochondrial location of the deficient acyl-CoA dehydrogenase by subfractionation of platelet mitochondria and, in contrast to the short-chain and medium-chain enzymes, which were localized in the soluble fraction, the majority of the acyl-CoA dehydrogenase activity with long-chain substrates was in the membrane fraction. These studies indicate that in humans, the predominant enzyme catalyzing the dehydrogenation of long-chain acyl-CoA esters is membrane-bound and that deficiency of this enzyme is a cause of muscle pain and rhabdomyolysis.

Acyl-CoA Dehydrogenase, Long-Chain↗
PubMed1994

Analysis of fatty acid oxidation intermediates in cultured fibroblasts to detect mitochondrial oxidation disorders.

We describe a method for the diagnosis of mitochondrial fatty acid oxidation disorders that is based on the analysis of acylcarnitine and acyl-coenzyme A (acyl-CoA) esters generated during fatty acid oxidation by permeabilized skin fibroblasts. This method requires only small amounts of cultured fibroblasts with minimal preparation, and no isolation of mitochondrial fractions is necessary. During oxidation of [U-14C]hexadecanoate, normal human fibroblasts from patients with fatty acid oxidation defects show a completely different pattern of intermediates, and in each case the observed profile reflects the site of the defect. The diagnosis and likely site of a mitochondrial fatty acid oxidation defect can be made readily from two 80-cm2 culture flasks of fibroblasts with this technique.

Acyl Coenzyme A↗
PubMed1993

Skeletal muscle mitochondrial beta-oxidation of dicarboxylates.

(1) The oxidation of [U-14C]hexadecanedionoyl-mono-CoA by rat skeletal muscle mitochondrial fractions is carnitine dependent and is inhibited by cyanide. (2) [U-14C]hexadecanedionoyl-mono-CoA was oxidised at a rate 8% of that of [U-14C]hexadecanoyl-CoA. (3) Oxidations were saturable and no substrate inhibition was observed. (4) We demonstrate the formation of dicarboxylyl-mono-CoA esters and the corresponding carnitine derivatives. (5) We conclude that, although skeletal muscle mitochondria are capable of the beta-oxidation of dicarboxylic acids, this is unlikely to be of great physiological significance.

Acyl Coenzyme A↗
PubMed1992

Intermediates of peroxisomal beta-oxidation of [U-14C]hexadecanedionoate. A study of the acyl-CoA esters which accumulate during peroxisomal beta-oxidation of [U-14C]hexadecanedionate and [U-14C]hexadecanedionoyl-mono-CoA.

1. The oxidation of [U-14C]hexadecanedionoyl-mono-CoA was stimulated by CoA, by carnitine in the absence of CoA and by the presence of an NAD(+)-regenerating system. 2. Substrate inhibition was observed with respect to [U-14C]hexadecanedionoyl-mono-CoA at concentrations greater than 35 microM. 3. Acetyl-CoA and the dicarboxyl-CoA esters of chain length C6-16 were detected by HPLC under standard incubation conditions. 4. In the absence of the NAD(+)-regenerating system, 2-enoyl-CoA and 3-hydroxacyl-CoA esters were detected. 5. In general, the peroxisomal beta-oxidation of dicarboxylates is very similar to that of monocarboxylates [Bartlett, K., Hovik, R., Eaton, S., Watmough, N. J. & Osmundsen, H. (1990) Biochem. J. 270, 175-180] except that chain shortening does not proceed beyond C6. 6. We conclude that the peroxisomal beta-oxidation of dicarboxylates is regulated by the redox state of the peroxisomal matrix and CoA availability.

Acetyl Coenzyme A↗
PubMed1992

Measurement of acyl-CoA dehydrogenase activity in cultured skin fibroblasts and blood platelets.

The measurement of acyl-CoA dehydrogenase activity is an essential part of the investigation of patients with suspected defects of fatty acid oxidation, and recently the organometallic oxidant ferricenium hexafluorophosphate has been introduced as an electron acceptor for these assays. However, we show that when medium-chain acyl-CoA dehydrogenase activity was measured in cultured skin fibroblasts and platelets from patients with proven defects of this enzyme, there was considerable residual enzyme activity when this electron acceptor was used. The ferricenium assay is not as specific as the anaerobic ETF-linked assay in the biochemical diagnosis of medium-chain acyl-CoA dehydrogenase deficiency in fibroblasts, and therefore is of limited clinical applicability in its present form.

Acyl-CoA Dehydrogenases↗
PubMed1992

Combined enzyme defect of mitochondrial fatty acid oxidation.

A young girl presented with recurrent episodes of muscle weakness culminating in a severe attack of generalized muscle weakness. In the muscle mitochondria from the patient there was an abnormal pattern of intermediates of beta-oxidation with an accumulation of 3-hydroxyacyl- and 2-enoyl-CoA and carnitine esters, and 3-oxoacylcarnitines. There was low activity of long-chain 3-hydroxyacyl-CoA dehydrogenase in mitochondria from all tissues. The activity of long-chain 2-enoyl-CoA hydratase was low in muscle mitochondria and 3-oxoacyl-CoA thiolase activity measured with 3-oxohexadecanoyl-CoA as substrate was low in fibroblast, muscle, and cardiac mitochondria but only partial deficiency was present when the activity was measured with 3-oxooctanoyl-CoA. The activity of the long-chain 3-hydroxyacyl-CoA dehydrogenase and long-chain 3-oxoacyl-CoA thiolase in fibroblasts from the patient's parents was intermediate between those of controls and the patient. The patient has a combined defect of the long-chain 3-hydroxyacyl-CoA dehydrogenase, long-chain 3-oxoacyl-CoA thiolase, and long-chain 2-enoyl-CoA hydratase which appears to be inherited in an autosomal recessive manner. This suggests there is a multifunctional enzyme catalyzing these activities in human mitochondria and that this enzyme is deficient in our patient.

3-Hydroxyacyl CoA Dehydrogenases↗
PubMed1991

Quantitation of acyl-CoA and acylcarnitine esters accumulated during abnormal mitochondrial fatty acid oxidation.

We have used radio-high pressure liquid chromatography to study the acyl-CoA ester intermediates and the acylcarnitines formed during mitochondrial fatty acid oxidation. During oxidation of [U-14C]hexadecanoate by normal human fibroblast mitochondria, only the saturated acyl-CoA and acylcarnitine esters can be detected, supporting the concept that the acyl-CoA dehydrogenase step is rate-limiting in mitochondrial beta-oxidation. Incubations of fibroblast mitochondria from patients with defects of beta-oxidation show an entirely different profile of intermediates. Mitochondria from patients with defects in electron transfer flavoprotein and electron transfer flavoprotein:ubiquinone oxido-reductase are associated with slow flux through beta-oxidation and accumulation of long chain acyl-CoA and acylcarnitine esters. Increased amounts of saturated medium chain acyl-CoA and acylcarnitine esters are detected in the incubations of mitochondria with medium chain acyl-CoA dehydrogenase deficiency, whereas long chain 3-hydroxyacyl-CoA dehydrogenase deficiency is associated with accumulation of long chain 3-hydroxyacyl- and 2-enoyl-CoA and carnitine esters. These studies show that the control strength at the site of the defective enzyme has increased. Radio-high pressure liquid chromatography analysis of intermediates of mitochondrial fatty acid oxidation is an important new technique to study the control, organization and defects of the enzymes of beta-oxidation.

3-Hydroxyacyl CoA Dehydrogenases↗
PubMed1991

Synthesis, characterisation and high-performance liquid chromatography of C6-C16 dicarboxylyl-mono-coenzyme A and -mono-carnitine esters.

The synthesis and purification of the mono-coenzyme A and mono-carnitine esters of the homologous series of straight-chain even-numbered dicarboxylic acids (C6-C16) is described. The corresponding 3-hydroxyacyl- and 2-enoyl-CoA esters were prepared enzymatically. A reversed-phase high-performance liquid chromatographic (HPLC) system for the analysis of the intact CoA esters is described and their chromatographic behaviour documented. Reversed-phase HPLC systems for the analysis of the 4-bromophenacyl derivatives of the dicarboxylyl-mono-carnitines and the 4-nitrobenzyl derivatives of the free acids are also described. Some preliminary studies of the metabolism of [U-14C]hexadecanedionoyl-mono-CoA by rat liver peroxisomes and rat skeletal muscle mitochondria are described illustrating the application of these methods.

Acyl Coenzyme A↗