Showing papers on "Acyl-CoA published in 2008"

The very-long-chain hydroxy fatty acyl-CoA dehydratase PASTICCINO2 is essential and limiting for plant development.

Abstract: Very-long-chain fatty acids (VLCFAs) are synthesized as acyl-CoAs by the endoplasmic reticulum-localized elongase multiprotein complex. Two Arabidopsis genes are putative homologues of the recently identified yeast 3-hydroxy-acyl-CoA dehydratase (PHS1), the third enzyme of the elongase complex. We showed that Arabidopsis PASTICCINO2 (PAS2) was able to restore phs1 cytokinesis defects and sphingolipid long chain base overaccumulation. Conversely, the expression of PHS1 was able to complement the developmental defects and the accumulation of long chain bases of the pas2–1 mutant. The pas2–1 mutant was characterized by a general reduction of VLCFA pools in seed storage triacylglycerols, cuticular waxes, and complex sphingolipids. Most strikingly, the defective elongation cycle resulted in the accumulation of 3-hydroxy-acyl-CoA intermediates, indicating premature termination of fatty acid elongation and confirming the role of PAS2 in this process. We demonstrated by in vivo bimolecular fluorescence complementation that PAS2 was specifically associated in the endoplasmic reticulum with the enoyl-CoA reductase CER10, the fourth enzyme of the elongase complex. Finally, complete loss of PAS2 function is embryo lethal, and the ectopic expression of PHS1 led to enhanced levels of VLCFAs associated with severe developmental defects. Altogether these results demonstrate that the plant 3-hydroxy-acyl-CoA dehydratase PASTICCINO2 is an essential and limiting enzyme in VLCFA synthesis but also that PAS2-derived VLCFA homeostasis is required for specific developmental processes.

The human peroxisomal ABC half transporter ALDP functions as a homodimer and accepts acyl-CoA esters

Abstract: Peroxisomes play a major role in human cellular lipid metabolism, including the β-oxidation of fatty acids. The most frequent peroxisomal disorder is X-linked adrenoleukodystrophy (X-ALD), which is caused by mutations in the ABCD1 gene. The protein involved, called ABCD1, or alternatively ALDP, is a member of the ATP-binding-cassette (ABC) transporter family and is located in the peroxisomal membrane. The biochemical hallmark of X-ALD is the accumulation of very long-chain fatty acids (VLCFAs), due to an impaired peroxisomal β-oxidation. Although this suggests a role of ALDP in VLCFA import, no experimental evidence is available to substantiate this. In the yeast Saccharomyces cerevisiae, peroxisomes are the exclusive site of fatty acid β-oxidation. Earlier work has shown that uptake of fatty acids into peroxisomes may occur via two routes, either as free fatty acids thus requiring intraperoxisomal activation into acyl-CoA esters or as long-chain acyl-CoA esters. The latter route involves the two peroxiso...

Short-and medium-chain carnitine acyltransferases and acyl-CoA thioesterases in mouse provide complementary systems for transport of β-oxidation products out of peroxisomes

Abstract: Peroxisomes metabolize a variety of lipids, acting as a chain-shortening system that produces acyl-CoAs of varying chain lengths, including acetyl-CoA and propionyl-CoA. It is, however, still largely unknown how β-oxidation products exit peroxisomes and where they are further metabolized. Peroxisomes contain carnitine acetyltransferase (CRAT) and carnitine octanoyltransferase (CROT) that produce carnitine esters for transport out of peroxisomes, together with recently characterized acyl-CoA thioesterases (ACOTs) that produce free fatty acids. Here we have performed tissue expression profiling of the short- and medium-chain carnitine acyltransferases Crat, Crot and the short- and medium-chain thioesterases (Acot12) and (Acot5), and show that they are largely expressed in different tissues, suggesting that they do not compete for the same substrates but rather provide complementary systems for transport of metabolites across the peroxisomal membrane. These data also explain earlier observed tissue differences in peroxisomal production of acetyl-CoA/acetyl-carnitine/acetate and underscores the differences in peroxisome function in various organs.

Fatty Acid Oxidation and Insulin Action: When Less Is More

Abstract: Type 2 diabetes is a disease of metabolic dysregulation involving impaired uptake and utilization of glucose, altered lipid metabolism, accumulation of various lipid species in the circulation and in tissues, and disruption of metabolic signaling pathways that regulate insulin secretion from pancreatic islet β-cells. Normal fuel homeostasis involves reciprocal regulation of glucose and lipid catabolism. Fundamental contributions to our understanding of the interplay between these two key groups of metabolic fuels came from the work of Randle (1), who demonstrated that increased rates of fatty acid oxidation in the fasted state lead to suppression of glucose oxidation and activation of gluconeogenesis, thereby preserving blood glucose for use by the brain and central nervous system. Conversely, the transition from the fasted to the fed state involves a coordinated shift from fatty acid to glucose oxidation. A key element in the latter switch, as elegantly demonstrated by the work of McGarry (2), is the glucose-induced rise in malonyl CoA, which inhibits fatty acid oxidation via direct binding to and allosteric inhibition of carnitine palmitoyltransferase-1 (CPT-1), the rate limiting enzyme for transport of cytosolic long-chain acyl CoA molecules into the mitochondria for oxidation. The multifaceted roles of malonyl CoA as a key glucose-derived metabolite, an allosteric inhibitor of fatty acid oxidation, and a biosynthetic precursor for fatty acid synthesis has led to a series of recent studies investigating the effects of manipulating this metabolite in various tissues. A bonus of such experiments is the opportunity to assess the physiological impact of enhanced or diminished fat oxidation in different cell types and in whole animals. In this issue of Diabetes …

Saturated fatty acids inhibit hepatic insulin action by modulating insulin receptor expression and post-receptor signalling.

Abstract: Free fatty acids (FFAs) are proposed to play a pathogenic role in both peripheral and hepatic insulin resistance. We have examined the effect of saturated FFA on insulin signalling (100 nM) in two hepatocyte cell lines. Fao hepatoma cells were treated with physiological concentrations of sodium palmitate (0.25 mM) (16:0) for 0.25-48 h. Palmitate decreased insulin receptor (IR) protein and mRNA expression in a dose- and time-dependent manner (35% decrease at 12 h). Palmitate also reduced insulin-stimulated IR and IRS-2 tyrosine phosphorylation, IRS-2-associated PI 3-kinase activity, and phosphorylation of Akt, p70 S6 kinase, GSK-3 and FOXO1A. Palmitate also inhibited insulin action in hepatocytes derived from wild-type IR (+/+) mice, but was ineffective in IR-deficient (-/-) cells. The effects of palmitate were reversed by triacsin C, an inhibitor of fatty acyl CoA synthases, indicating that palmitoyl CoA ester formation is critical. Neither the non-metabolized bromopalmitate alone nor the medium chain fatty acid octanoate (8:0) produced similar effects. However, the CPT-1 inhibitor (+/-)-etomoxir and bromopalmitate (in molar excess) reversed the effects of palmitate. Thus, the inhibition of insulin signalling by palmitate in hepatoma cells is dependent upon oxidation of fatty acyl-CoA species and requires intact insulin receptor expression.

The Nudix Hydrolase 7 is an Acyl-CoA Diphosphatase Involved in Regulating Peroxisomal Coenzyme A Homeostasis

Abstract: Coenzyme A (CoASH) is an obligate cofactor for lipids undergoing beta-oxidation in peroxisomes. Although the peroxisomal membrane appears to be impermeable to CoASH, peroxisomes contain their own pool of CoASH. It is believed that CoASH enters peroxisomes as acyl-CoAs, but it is not known how this pool is regulated. The mouse nudix hydrolase 7 (NUDT7alpha) was previously identified in peroxisomes as a CoA-diphosphatase, and therefore suggested to be involved in regulation of peroxisomal CoASH levels. Here we show that mouse NUDT7alpha mainly acts as an acyl-CoA diphosphatase, with highest activity towards medium-chain acyl-CoAs, and much lower activity with CoASH. Nudt7alpha mRNA is highly expressed in liver, brown adipose tissue and heart, similar to enzymes involved in peroxisomal lipid degradation. Nudt7alpha mRNA is down-regulated by Wy-14,643, a peroxisome proliferator-activated receptor alpha (PPARalpha) ligand, in a PPARalpha-dependent manner in mouse liver. In highly purified peroxisomes, nudix hydrolase activity is highest with C(6)-CoA and is decreased by fibrate treatment. Under certain conditions, such as treatment with peroxisome proliferators or fasting, an increase in peroxisomal CoASH levels has been reported, which is in line with a decreased expression/activity of NUDT7alpha. Taken together these data suggest that NUDT7alpha function is tightly linked to peroxisomal CoASH/acyl-CoA homeostasis.

Crocetin improves the insulin resistance induced by high-fat diet in rats

Abstract: Background and purpose: The amelioration of insulin resistance by treatment with crocetin is closely related to the hypolipidaemic effect. The present study is designed to clarify the insulin-sensitizing mechanism of crocetin by elucidating the mechanism of regulation of lipid metabolism by crocetin. Experimental approach: Rats given a high-fat diet were treated with crocetin for 6 weeks before hyperinsulinaemic–euglycaemic clamp. 14C-palmitate was used as tracer to track the fate of non-esterified fatty acids or as substrate to measure β-oxidation rate. Triglyceride clearance in plasma and lipoprotein lipase activity in tissues were tested. Content of lipids in plasma and tissues was determined. Real-time PCR was used to assay the level of mRNA from genes involved in non-esterified fatty acid and triglyceride uptake and oxidation. Key results: Crocetin prevented high-fat-diet induced insulin resistance (increased clamp glucose infusion rate), raised hepatic non-esterified fatty acid uptake and oxidation, accelerated triglyceride clearance in plasma, enhanced lipoprotein lipase activity in liver, and reduced the accumulation of detrimental lipids (DAG and long-chain acyl CoA) in liver and muscle. Genes involved in hepatic lipid metabolism which are regulated by peroxisome proliferator-activated receptor-α, were modulated to accelerate lipid uptake and oxidation. Conclusions and implications: Through regulating genes involved in lipid metabolism, crocetin accelerated hepatic uptake and oxidation of non-esterified fatty acid and triglyceride, and reduced lipid availability to muscle, thus decreasing lipid accumulation in muscle and liver, and consequently improving sensitivity to insulin. British Journal of Pharmacology (2008) 154, 1016–1024; doi:10.1038/bjp.2008.160; published online 12 May 2008

Long chain fatty acyl-CoA modulation of H2O2 release at mitochondrial complex I

Abstract: Complex I is responsible for most of the mitochondrial H2O2 release, low during the oxidation of the NAD linked substrates and high during succinate oxidation, via reverse electron flow. This H2O2 production appear physiological since it occurs at submillimolar concentrations of succinate also in the presence of NAD substrates in heart (present work) and rat brain mitochondria (Zoccarato et al., Biochem J, 406:125–129, 2007). Long chain fatty acyl-CoAs, but not fatty acids, act as strong inhibitors of succinate dependent H2O2 release. The inhibitory effect of acyl-CoAs is independent of their oxidation, being relieved by carnitine and unaffected or potentiated by malonyl-CoA. The inhibition appears to depend on the unbound form since the acyl-CoA effect decreases at BSA concentrations higher than 2 mg/ml; it is not dependent on ΔpH or Δp and could depend on the inhibition of reverse electron transfer at complex I, since palmitoyl-CoA inhibits the succinate dependent NAD(P) or acetoacetate reduction.

Modulation of hSlo BK current inactivation by fatty acid esters of CoA.

Abstract: Lipid metabolism influences membrane proteins, including ion channels, in health and disease. Fatty acid esters of CoA are important intermediates in fatty acid metabolism and lipid biosynthesis. In the present study, we examined the effect of acyl-CoAs on hSlo BK currents. Arachidonoyl-CoA (C20-CoA) induced β2-dependent inhibition of hSlo-α current when applied intracellularly but not extracellularly. This action was also mimicked by other long-chain acyl-CoAs such as oleoyl-CoA (C18-CoA) and palmitoyl-CoA (C16-CoA), but not acetyl-CoA (C2-CoA, shorter chain), suggesting that the length of acyl chains, rather than CoA headgroups, is critical. When hSlo-α inactivation was induced by a free synthetic cationic β2 NH2-terminus inactivation ball peptide, long-chain acyl-CoAs inhibited hSlo-α current and facilitated inactivation. The precursor fatty acids also facilitated the ball peptide-induced inactivation in a chain length-dependent manner, whereas sphingosine (positively charged) slowed this inactivation. When the β2-induced inactivation was compared with that of the ball peptide, there was a negative shift in the steady state inactivation, slower recovery, and a reduced voltage-dependence of inactivation onset. These data suggest that electrostatic interactions with the cytosolic inactivation domain of β2 mediate acyl-CoA modulation of BK currents. BK channel inactivation may be a specific target for lipid modulation in physiological and pathophysiological conditions.

Mycobacterium tuberculosis β-Ketoacyl Acyl Carrier Protein Synthase III (mtFabH) Assay: Principles and Method

Abstract: Fatty acid biosynthesis is one of the relatively newer targets in antibacterial drug discovery. The presence of distinct fatty acid synthases (FAS) in mammals and bacteria and the fact that most bacterial FAS enzymes are essential for viability make this a very attractive antimicrobial drug target. The enzyme beta-ketoacyl ACP synthase (KASIII or FabH) is the key enzyme that initiates fatty acid biosynthesis in a type II dissociated FAS. This enzyme catalyzes the condensation of acyl CoA and malonyl ACP (acyl carrier protein) to form a beta-ketoacyl ACP product, which is further processed to form mature fatty acids that are involved in various essential cellular processes and structures like phospholipid biosynthesis, cell wall formation, etc. Herein we describe a new assay for the Mycobacterium tuberculosis FabH (mtFabH) enzyme involved in a key initiation step in the synthesis of mycolic acids, which are an integral component of the cell wall. The assay eliminates the need for the cumbersome washing steps or specialty scintillation proximity assay beads and the preparation of acyl carrier proteins required in other assay formats. This discontinuous assay involves the reduction of radiolabled long-chain beta-ketoacyl CoA product to its dihydroxy derivative, which partitions into a nonpolar phase for quantitation, while the reduced radiolabeled substrate derivative remains in the aqueous phase.

Fat metabolism promoter

Abstract: PROBLEM TO BE SOLVED: To provide a fat metabolism promoter containing a new ingredient metabolizing a fat as an active ingredient and to provide a fat metabolism promoter having a new ingredient promoting expression of enzyme gene of carnitine palmitoyltransferase and acyl CoA oxydase which are each an enzyme which becomes a rate-controlling step of fat metabolism and a peroxisome proliferators activated receptor (PPAR) as an active ingredient. SOLUTION: The fat metabolism promoter, a liver fat metabolism promoter, a muscle fat metabolism promoter, a liver carnitine palmitoyltransferase activating agent, a liver acyl CoA oxydase activating agent and a liver PPAR activating agent, a muscle carnitine palmitoyltransferase activating agent and a PPAR activating agent each contains an α-lipoic acid. COPYRIGHT: (C)2008,JPO&INPIT

Retraction: Protective Effect of PGC-1 on Lipid Overload-induced Apoptosis in Vascular Endothelial Cell

Abstract: Background: Fatty acids contribute to endothelial cell dysfunction and apoptosis by inducing accumulation of long chain fatty acyl CoA (LCAC), which increases oxidative stress in vascular endothelial cells. Forced expression of PGC-1 was shown to induce mitochondrial biogenesis and to control expression of mitochondrial enzymes involved in fatty acid oxidation. This study was undertaken to test the hypothesis that PGC-1 overexpression could prevent endothelial cell apoptosis by enhancing fatty acid oxidation and relieving oxidative stress in vascular endothelium. Methods: Adenoviruses containing human PGC-1 (Ad-PGC-1) and -galactosidase (Ad--gal) were transfected to confluent human aortic endothelial cells (HAECs). To investigate the effect of adenoviral PGC-1 gene transfer on apoptosis, combined treatment of linoleic acid (LA), an unsaturated fatty acid, was performed. Results: PGC-1 overexpression inhibited the increase in ROS production and apoptosis of HAECs induced by LA. Also, PGC-1 led to a significant increase in fatty acid oxidation and decrease in triglyceride content in HAECs. LA caused the decrease of adenine nucleotide translocase (ANT) activity and transient mitochondrial hyperpolarization, which was followed by depolarization. PGC-1 overexpression prevented these processes. Conclusion: In summary, PGC-1 overexpression inhibited mitochondrial dysfunction and apoptosis by facilitating fatty acid oxidation and protecting against the damage from oxidative stress in HAECs. The data collectively suggest that the regulation of intracellular PGC-1 expression might play a critical role in preventing atherosclerosis.

Correlation between peroxisome proliferation and upregulation of cytochrome P450 CYP4A and peroxisomal beta-oxidation fatty acyl CoA oxidases (AOX) in the koala (Phascolarctos cinereus)

Abstract: Peroxisomes are membrane bound cytoplasmic organelles that are involved in lipid metabolism and other biological functions. In rat and mouse, profound xenobiotic-induced peroxisome proliferation has been reported, with a marked increase in number and size of peroxisomes in liver parenchymal cells and induction of lipid metabolising enzymes, in particular cytochrome P450 CYP4A and peroxisomal β-oxidation palmitoyl CoA oxidases (AOX1). The present study investigates whether the previously observed higher hepatic CYP4A and AOX1 expression in the koala (Phascolarctos cinereus), a unique Australian marsupial, compared with rat and human is associated with peroxisome proliferation. Visualisation and quantification of peroxisomes were performed on liver samples from three koalas utilising transmission electron microscopy, with rat and bandicoot livers being used for comparative purposes. Numerous catalase positive peroxisomes, which clearly stand out by their black single membrane globular structures, were detected in all test ultra-thin sections from koala livers. A higher average number of peroxisomes per hepatocyte was observed for the koala, an obligate eucalyptus feeder, compared with non-eucalyptus feeders rat and bandicoot. No species differences in the average size of peroxisomes were detected. This is the first morphological study examining hepatic peroxisomes in an Australian marsupial. The results suggested that dietary eucalyptus constituents might possess peroxisome proliferating activities.

Method for acyltransferase reaction using acyl coenzyme a

Abstract: The present invention relates to a method for acyltransferase reaction in which an acyl group of acyl coenzyme A is transferred to an acyl group receptor characterized in that the reaction is carried out by production and/or reproduction of acyl coenzyme A from coenzyme A in a reaction system by a chemical thioester exchange reaction with acylthioester. The present invention, wherein expensive acyl CoA is reproduced nonenzymatically in a reaction system, enables to continuously carry out acyltransferase reaction only by putting a small amount of acyl CoA with a donor and a receptor of an acyl group into a system. Accordingly, the method of the present invention can be applied to an industrial production method of various kinds of compounds including useful biological molecules and synthesis of polymers such as polyester.