Showing papers on "Acyl-CoA published in 2012"

The hexanoyl-CoA precursor for cannabinoid biosynthesis is formed by an acyl-activating enzyme in Cannabis sativa trichomes.

Abstract: *SUMMARY The psychoactive and analgesic cannabinoids (e.g. D 9 -tetrahydrocannabinol (THC)) in Cannabis sativa are formed from the short-chain fatty acyl-coenzyme A (CoA) precursor hexanoyl-CoA. Cannabinoids are synthesized in glandular trichomes present mainly on female flowers. We quantified hexanoyl-CoA using LC-MS/MS and found levels of 15.5 pmol g )1 fresh weight in female hemp flowers with lower amounts in leaves, stems and roots. This pattern parallels the accumulation of the end-product cannabinoid, cannabidiolic acid (CBDA). To search for the acyl-activating enzyme (AAE) that synthesizes hexanoyl-CoA from hexanoate, we analyzed the transcriptome of isolated glandular trichomes. We identified 11 unigenes that encoded putative AAEs including CsAAE1, which shows high transcript abundance in glandular trichomes. In vitro assays showed that recombinant CsAAE1 activates hexanoate and other short- and medium-chained fatty acids. This activity and the trichome-specific expression of CsAAE1 suggest that it is the hexanoyl-CoA synthetase that supplies the cannabinoid pathway. CsAAE3 encodes a peroxisomal enzyme that activates a variety of fatty acid substrates including hexanoate. Although phylogenetic analysis showed that CsAAE1 groups with peroxisomal AAEs, it lacked a peroxisome targeting sequence 1 (PTS1) and localized to the cytoplasm. We suggest that CsAAE1 may have been recruited to the cannabinoid pathway through the loss of its PTS1, thereby redirecting it to the cytoplasm. To probe the origin of hexanoate, we analyzed the trichome expressed sequence tag (EST) dataset for enzymes of fatty acid metabolism. The high abundance of transcripts that encode desaturases and a lipoxygenase suggests that hexanoate may be formed through a pathway that involves the oxygenation and breakdown of unsaturated fatty acids.

Characterization of D-3-hydroxybutyrylcarnitine (ketocarnitine): an identified ketosis-induced metabolite.

Abstract: Hydroxybutyrylcarnitine (HB-carnitine) is a metabolite that has been associated with insulin resistance and type 2 diabetes mellitus. It is currently unknown whether HB-carnitine can be produced from D-3-hydroxybutyrate (D-3HB), a ketone body; but its formation from L-3-HB-CoA, a fatty acid β-oxidation intermediate, is well established. We aimed to assess which stereoisomers of 3-HB-carnitine are present in vivo. Ketosis and increased fatty acid oxidation were induced in 12 lean healthy men by a 38-hour fasting period. The D-3HB kinetics (stable isotope technique) and stereoisomers of muscle 3-HB-carnitine (high-performance liquid chromatography/ultra-performance liquid chromatography-tandem mass spectrometry) were measured. Muscle D-3HB-carnitine content was much higher compared with L-3HB-carnitine. In addition, muscle D-3HB-carnitine correlated significantly with D-3-HB production. Following the finding that a ketone body can be converted into a carnitine ester in vivo, we show in vitro that D-3-HB can be converted into HB-carnitine (ketocarnitine) via an acyl-CoA synthetase reaction in several tissues including human muscle. During fasting, HB-carnitine in muscle is derived mainly from the ketone body D-3HB. The role of D-3HB-carnitine synthesis in metabolism remains to be elucidated.

Pseudomonas aeruginosa Directly Shunts β-Oxidation Degradation Intermediates into De Novo Fatty Acid Biosynthesis

Abstract: We identified the fatty acid synthesis (FAS) initiation enzyme in Pseudomonas aeruginosa as FabY, a β-ketoacyl synthase KASI/II domain-containing enzyme that condenses acetyl coenzyme A (acetyl-CoA) with malonyl-acyl carrier protein (ACP) to make the FAS primer β-acetoacetyl-ACP in the accompanying article (Y. Yuan, M. Sachdeva, J. A. Leeds, and T. C. Meredith, J. Bacteriol. 194:5171-5184, 2012). Herein, we show that growth defects stemming from deletion of fabY can be suppressed by supplementation of the growth media with exogenous decanoate fatty acid, suggesting a compensatory mechanism. Fatty acids eight carbons or longer rescue growth by generating acyl coenzyme A (acyl-CoA) thioester β-oxidation degradation intermediates that are shunted into FAS downstream of FabY. Using a set of perdeuterated fatty acid feeding experiments, we show that the open reading frame PA3286 in P. aeruginosa PAO1 intercepts C8-CoA by condensation with malonyl-ACP to make the FAS intermediate β-keto decanoyl-ACP. This key intermediate can then be extended to supply all of the cellular fatty acid needs, including both unsaturated and saturated fatty acids, along with the 3-hydroxyl fatty acid acyl groups of lipopolysaccharide. Heterologous PA3286 expression in Escherichia coli likewise established the fatty acid shunt, and characterization of recombinant β-keto acyl synthase enzyme activity confirmed in vitro substrate specificity for medium-chain-length acyl CoA thioester acceptors. The potential for the PA3286 shunt in P. aeruginosa to curtail the efficacy of inhibitors targeting FabY, an enzyme required for FAS initiation in the absence of exogenous fatty acids, is discussed.

Myocardial Ischemia and Lipid Metabolism

Abstract: Lipid Metabolism in the Myocardium.- Overview of Lipid Metabolism.- Factors Influencing the Carnitine-Dependent Oxidation of Fatty Acids in the Heart.- Localization and Function of Lipases and Their Reaction Products in Rat Heart.- Ultrastructural Localization of Lipids in Myocardial Membranes.- Lipid Induced Membrane Abnormalities.- Phospholipase-Induced Abnormalities in the Sarcolemma.- A Surface Charge Hypothesis for the Actions of Palmitylcarnitine on the Kinetics of Excitatory Ionic Currents in Heart.- Modulation of Membrane Function by Lipid Intermediates: A Possible Role in Myocardial Ischemia.- Fatty Acid Effects on Sarcoplasmic Reticulum Function In Vitro.- Ischemia and Lipid-Induced Changes in Myocardial Function.- Factors Influencing the Metabolic and Functional Alterations Induced by Ischemia and Reperfusion.- Factors that Influence Myocardial Levels of Long-Chain Acyl CoA and Acyl Carnitine.- Are Tissue Non-Esterified Fatty Acids (NEFA) Involved in the Impairment of Biochemical and Mechanical Processes during Acute Regional Ischemia in the Heart.- Effects of Myocardial Ischemia and Long Chain Acyl CoA on Mitochondrial Adenine Nucleotide Translocator.- Interventions Used to Modify Lipid-Induced Abnormalities in the Heart.- Fatty Acid and Carnitine-Linked Abnormalities during Ischemia and Cardiomyopathy.- Consequences of Fatty Acid Excess in Ischemic Myocardium and Effects of Therapeutic Interventions.- Membrane Phospholipid Metabolism during Myocardial Ischemia: Mechanisms of Accumulation of Unesterified Arachidonate.- Phospholipase and Ischemic Damage: Possibilities of Interventions.- Importance of Lipid Metabolism in Man.- Cardiac Perfusion, Past and Present.- Clinical Relevance of Free Fatty Acid Excess.- Iodine-123 Phenylpentadecanoic Acid: Detection of Acute Myocardial Infarction in Anesthetized Dogs.- Free Fatty Acid, Catecholamines and Arrhythmias in Man.- Conclusions.- Contributors.

Molecular and functional analysis of three fatty acyl-CoA reductases with distinct substrate specificities in copepod Calanus finmarchicus.

Abstract: The marine copepod Calanus finmarchicus constitutes the substantial amount of biomass in the Arctic and Northern seas. It is unique in that this small crustacean accumulates a high level of wax esters as carbon storage which is mainly comprised of 20:1n-9 and 22:1n-11 alcohols (Alc) linked with various kinds of fatty acids, including n-3 polyunsaturated fatty acids. The absence of 20:1n-9 Alc and 22:1n-11 Alc in diatoms and dinoflagellates, the primary food sources of copepods, suggests the existence of de novo biosynthesis of fatty alcohols in C. finmarchinus. Here, we report identification of three genes, CfFAR1, CfFAR2, and CfFAR3, coding for fatty acyl-CoA reductases involved in the conversion of various fatty acyl-CoAs to their corresponding alcohols. Functional characterization of these genes in yeast indicated that CfFAR1 could use a wide range of saturated fatty acids from C18 to C26 as substrates, CfFAR2 had a narrow range of substrates with only very-long-chain saturated fatty acid 24:0 and 26:0, while CfFAR3 was active towards both saturated (16:0 and 18:0) and unsaturated (18:1 and 20:1) fatty acids producing corresponding alcohols. This finding suggested that these three fatty acyl-CoA reductases are likely responsible for de novo synthesis of a series of fatty alcohol moieties of wax esters in C. finmarchicus.

Differential Effect of Fatty Acids in Nervous Control of Energy Balance

Abstract: Energy homeostasis is kept through a complex interplay of nutritional, neuronal and hormonal inputs that are integrated at the level of the central nervous system (CNS). A disruption of this regulation gives rise to life-threatening conditions that include obesity and type-2 diabetes, pathologies that are strongly linked epidemiologically and experimentally. The hypothalamus is a key integrator of nutrient-induced signals of hunger and satiety, crucial for processing information regarding energy stores and food availability. Much effort has been focused on the identification of hypothalamic pathways that control food intake but, until now, little attention has been given to a potential role for the hypothalamus in direct control of glucose homeostasis and nergy balance. Recent studies have cast a new light on the role of the CNS in regulating peripheral glucose via a hypothalamic fatty acid (FA)-sensing device that detects nutrient availability and relays, through the autonomic nervous system, a negative feedback signal on food intake, insulin sensitivity and insulin secretion. Indeed, accumulating evidences suggest that FA are used in specific areas of CNS not as nutrients, but as cellular messengers which inform “FA sensitive neurons” about the energy status of the whole body (Blouet & Schwartz, 2010; Migrenne et al., 2006; Migrenne et al., 2011). Thus it has been described that up to 70% of hypothalamic arcuate nucleus (ARC) and ventromedian nucleus (VMN) neurons are either excited or inhibited by long chain fatty acids such as oleic acid (Jo et al., 2009; Le Foll et al., 2009; Migrenne et al., 2011). Within the VMN, 90% of the glucosensing neurons also have their activity altered by FA. In a large percentage of these neurons, glucose and FA have opposing effects on neuronal activity, much as they do on intracellular metabolism in many other cells (Randle et al., 1994). Neuronal FA sensing mechanisms include activation of the KATP channel by long chain fatty acid acyl CoA (Gribble et al., 1998) or inactivation by generation of ATP or reactive oxygen species during mitochondrial ┚-oxidation (Jo et al., 2009; Le Foll et al., 2009; Migrenne et al., 2011; Wang et al., 2006). Many fatty acid sensing neurons are activated by interaction of long chain fatty acids with the fatty acid transporter/receptor, FAT/CD36, presumably by activation of store-operated calcium channels by a mechanism that is independent of fatty acid metabolism (Jo et al., 2009). Importantly, most neurons utilize FA primarily for membrane production rather than as a metabolic substrate (Rapoport et al., 2001; Smith & Nagura, 2001) and only nanomolar concentrations of fatty acid are required to

Impact of specific long chain acyl CoA synthetases on plant development

Abstract: Fatty acids are essential components of cellular life. However, in their natural form they cannot enter metabolism but in fact they need enzymatic activation by coenzyme A to become metabolically accessible. One enzyme class catalyzing such a reaction is called long chain acyl-CoA synthetases (LACS). Although the chemical aspects of this reaction are well understood the biological impact remained largely unknown, since pronounced phenotypes of LACS mutants have not been reported so far. However, in this project it has been shown that deletion of these activities can lead to severe phenotypes affecting processes throughout the whole plant life cycle. The observations were made by the establishment of a LACS mutant collection with dozen of double and triple knock-out lines. Some lines showed reduced or even absent fertility and reduced surface wax levels caused by a reduced synthesis of very long chain lipids. Other mutants showed light dependent phenotypes resulting in severely altered plant morphology. These modifications were most likely due to perturbations of the lipid metabolism. In this context, a reduced flux between prokaryotic and eukaryotic pathway is assumed based on radiolabeled flux analysis. In addition, embryo development and storage lipid synthesis were reduced in some lines indicating important roles of specific LACS enzymes also in these processes. All these phenotypes are caused by specific changes in various parts of lipid metabolism, suggesting that fatty acids require repeated activation, after being transported out of the chloroplast. Evidences for such processes have been found in surface wax synthesis, storage lipid synthesis, fatty acid degradation and in part also for eukaryotic lipid synthesis. Deactivation of acyl-CoA and subsequent reactivation of the free fatty acid seem to be contra productive; however, it might allow a better more efficient regulation of the connected metabolic pathways or transport processes through membranes. Furthermore, the results are suggesting the existence of different acyl CoA pools for specific pathways. Such different pools might also be the reason for the existence of nine LACS isogenes in Arabidopsis.