Acyl Coenzyme A Synthetase Research Articles

Roles of acyl‐coenzyme A synthetase family members in airway epithelia cells

AbstractBackgroundThe fundamental initial step in the metabolic process of fatty acids (FA) is facilitated by Acyl‐coenzyme A synthetase (ACS). ACS enables FAs to engage in both anabolic and catabolic pathways by creating thioester bonds with CoA, thus triggering their activation. This paper demonstrated the effect of in vitro cells on the ACS family of key enzymes in FA metabolism in response to inflammatory or lipid stimulating factors and provided insight into the relevant metabolic mechanisms.MethodsThe expression levels of 27 ACS family members were assessed across various lung epithelial cell lines, such as HBE, SPC‐1A, or NCI‐H460, in response to external challenges including lipopolysaccharide (LPS), lysophosphatidylcholine (lysoPC), cigarette smoking extract (CSE), cholesterol, and interference with the ACSL5 gene in response to FASN, CPT1A inhibitors.ResultsOur data showed that ACSF2 was highly expressed in SPC‐A1 and NCI‐H460 cells compared with HBE cell, while ACSL4 and ACSVL4 were lower expressed. The 27 ACS family genes associated with FA metabolism varied under several different exogenous stimuli.

Acute Testosterone Deficiency Alters Adipose Tissue Fatty Acid Storage.

Although the long-term effects of testosterone on adipose tissue lipid metabolism in men have been defined, the short-term regulation of these effects is not well understood. We examined the effects of acute testosterone withdrawal on subcutaneous abdominal and femoral adipose tissue fatty acid (FA) storage and cellular mechanisms. This was a prospective, randomized trial. Mayo Clinic Clinical Research Unit. Thirty-two male volunteers ages 18 to 50 participated in these studies. Volunteers were randomized to receive (1) no treatment (control), (2) injections (7.5 mg) of Lupron®, or (3) Lupron and testosterone (L+T) replacement for 49 days, resulting in 4 weeks of sex steroid suppression in the Lupron group. We measured body composition, fat cell size, adipose tissue meal FA and direct free FA storage, lipoprotein lipase (LPL), acyl coenzyme A synthetase (ACS), diacylglycerol acyltransferase activities, and CD36 content. Compared with control and L+T groups, acute testosterone deficiency resulted in greater femoral adipose tissue meal FA storage rates, fasting and fed LPL activity, and ACS activity. These results suggest that in men, testosterone plays a tonic role in restraining FA storage in femoral adipose tissue via suppression of LPL and ACS activities. FA storage mechanisms in men appear sensitive to short-term changes in testosterone concentrations.

Systematic Analysis of the 4-Coumarate:Coenzyme A Ligase (4CL) Related Genes and Expression Profiling during Fruit Development in the Chinese Pear.

In plants, 4-coumarate:coenzyme A ligases (4CLs), comprising some of the adenylate-forming enzymes, are key enzymes involved in regulating lignin metabolism and the biosynthesis of flavonoids and other secondary metabolites. Although several 4CL-related proteins were shown to play roles in secondary metabolism, no comprehensive study on 4CL-related genes in the pear and other Rosaceae species has been reported. In this study, we identified 4CL-related genes in the apple, peach, yangmei, and pear genomes using DNATOOLS software and inferred their evolutionary relationships using phylogenetic analysis, collinearity analysis, conserved motif analysis, and structure analysis. A total of 149 4CL-related genes in four Rosaceous species (pear, apple, peach, and yangmei) were identified, with 30 members in the pear. We explored the functions of several 4CL and acyl-coenzyme A synthetase (ACS) genes during the development of pear fruit by quantitative real-time PCR (qRT-PCR). We found that duplication events had occurred in the 30 4CL-related genes in the pear. These duplicated 4CL-related genes are distributed unevenly across all pear chromosomes except chromosomes 4, 8, 11, and 12. The results of this study provide a basis for further investigation of both the functions and evolutionary history of 4CL-related genes.

Amelioration of Cryptosporidium parvum Infection In Vitro and In Vivo by Targeting Parasite Fatty Acyl-Coenzyme A Synthetases

Cryptosporidium is emerging as 1 of the 4 leading diarrheal pathogens in children in developing countries. Its infections in patients with AIDS can be fatal, whereas fully effective treatments are unavailable. The major goal of this study is to explore parasite fatty acyl-coenzyme A synthetase (ACS) as a novel drug target. A colorimetric assay was developed to evaluate biochemical features and inhibitory kinetics of Cryptosporidium parvum ACSs using recombinant proteins. Anticryptosporidial efficacies of the ACS inhibitor triacsin C were evaluated both in vitro and in vivo. Cryptosporidium ACSs displayed substrate preference toward long-chain fatty acids. The activity of parasite ACSs could be specifically inhibited by triacsin C with the inhibition constant Ki in the nanomolar range. Triacsin C was highly effective against C. parvum growth in vitro (median inhibitory concentration, 136 nmol/L). Most importantly, triacsin C effectively reduced parasite oocyst production up to 88.1% with no apparent toxicity when administered to Cryptosporidium-infected interleukin 12 knockout mice at 8-15 mg/kg/d for 1 week. The findings of this study not only validated Cryptosporidium ACS (and related acyl-[acyl-carrier-protein]-ligases) as pharmacological targets but also indicate that triacsin C and analogues can be explored as potential new therapeutics against the virtually untreatable cryptosporidial infection in immunocompromised patients.

Effects of Changes of Diet from High‐fat to Low‐fat on Hepatic mRNA Levels of ACS, CPT‐1, and ACC

AimsThe purpose of this study was to investigate the effects of dietary changes from high‐fat diet (HFD, 45% of total calories from fat) to low‐fat diet (LFD, 10% of total calories from fat) on hepatic mRNA levels of acyl coenzyme A synthetase(ACS), carnitine palmitoyltransferase‐1(CPT‐1), and acetyl coenzyme A carboxylase(ACC) as well as on serum and hepatic lipids in mice.MethodsFive weeks‐old male C57BL/6 mice were fed high‐fat diet for 9 weeks then switched to a low‐fat diet or continued the high‐fat diet for 3 weeks. Body weight, serum and hepatic lipid levels were measured. Expressions of ACS, CPT‐1, and ACC mRNA were also analyzed.ResultsThe results showed that after the high‐fat diets were switched to low fat diets, body weight, epididymal and retroperitoneal fats were significantly reduced (p<0.05). The levels of serum and hepatic total cholesterol were not different between the two groups however, serum TG levels of high‐fat diet followed by low‐fat diet group were significantly lower than those of high‐fat diet group (p<0.05). Hepatic mRNA levels of CPT‐1 and ACC were higher significantly in the group of high‐fat diet followed by low‐fat diet (p<0.05) however mRNA levels of ACS were not different between the two groups.ConclusionsChanges in the composition of dietary fat from high to low decreased significantly body fats and the serum TGs. Furthermore, changes of dietary fat composition affected differently the hepatic mRNA levels of ACS, CPT‐1 and ACC.

Impaired intrinsic chiral inversion activity of ibuprofen in rats with adjuvant-induced arthritis

1. The effects of adjuvant-induced arthritis on the chiral inversion of ‘profens’, a type of non-steroidal anti-inflammatory drug, have hardly been investigated. The authors investigated the effects of adjuvant-induced arthritis on the chiral inversion of ibuprofen using freshly isolated rat hepatocytes.2. S- or R-ibuprofen was incubated with hepatocytes isolated from control and adjuvant-induced arthritis rats in the absence of the serum. In the hepatocyte system the chiral inversion rate constant of R- to S-ibuprofen and the metabolic rate constants of both enantiomers in adjuvant-induced arthritis rats were significantly decreased to about 64–80% of the corresponding values in control rats. In contrast, the addition of serum from each group to the corresponding hepatocyte medium resulted in no significant differences in these rate constants between control and adjuvant-induced arthritis rats.3. With regard to chiral inversion enzymes, adjuvant-induced arthritis decreased the messenger RNA levels of acyl-coenzyme A synthetase (ACS) isoforms, but not 2-arylpropionyl-CoA epimerase, compared with control rats.4. Chiral inversion of R- to S-ibuprofen was inhibited by triacsin C, a specific inhibitor of ACS1.5. The results suggest that adjuvant-induced arthritis induces down-regulation of ACS enzymes involved in chiral inversion of R- to S-ibuprofen in rats.

Ultra-fast simultaneous detection of obesity-related coenzymes in mice using microchip electrophoresis with a LIF detector

Hepatic acyl-coenzyme A synthetase (ACS), carnitine palmitoyltransferase-I (CPT-I) and acetyl coenzyme A carboxylase (ACC) are coenzymes associated with the genetic type of obesity in animal models. This paper reports the use of microchip electrophoresis (ME) with a laser-induced fluorescence (LIF) detector based on a reverse transcriptase-polymerase chain reaction (RT-PCR) to detect the amplified DNA fragments of these coenzymes (ACS, CPT-I and ACC) in the mRNA extracted from mice. DNA fragments ranging from 50 to 2652 bp were well resolved using this procedure with a running buffer (1× TBE), 0.5% polyvinylpyrrolidone ( M r 1,000,000) as the coating gel and 0.7% polyethyleneoxide ( M r 8,000,000) as the sieving gel at pH 8.30. The separation of the three RT-PCR products was achieved by ME in a single-run within 17 s using programmed field strength gradients (PFSG) (470 V cm −1 for 9 s, 205.8 V cm −1 for 2 s, 411.6 V cm −1 for 4 s, 117.6 V cm −1 for 2 s and 470.4 V cm −1 for 8 s). The ME-PFSG method was found to be 4 times faster than the method using a constant field and 138 times faster than slab gel electrophoresis. Moreover, the amplified RT-PCR products of the obesity-related coenzymes in C57BL/6J mice were analyzed using only sub-micro liter samples.

Effect of Genistein with Carnitine Administration on Lipid Parameters and Obesity in C57Bl/6J Mice Fed a High-Fat Diet

Soy products are mainly composed of proteins, phytochemicals such as isoflavones, soy lipids, and carbohydrates. It is unclear whether an individual component alone or a combined effect of multiple bioactive compounds contributes to the beneficial properties of soy. We investigated the effect of dietary genistein (the principal soy isoflavone) alone and combined with L-carnitine to evaluate possible synergistic effects on the intentionally induced prediabetic state characterized by insulin resistance and obesity in C57Bl/6J mice fed a high-fat diet (HD). In the HD-alone group, abdominal and back fat relative to total body weight were significantly higher compared with other groups including those fed normal diet (ND). Among the HD groups, final weight gains of the HD plus genistein (HD+G) and HD plus genistein plus L-carnitine (HD+G+C) groups were lower compared with that of the control (HD-alone). Especially in liver, the results showed that genistein with carnitine transcriptionally up-regulated expressions of acyl-coenzyme A synthetase (ACS) and carnitine palmitoyltransferase-I (CPT-I) by approximately 50% and 40%, respectively, compared with genistein alone. However, the up-regulation of CPT-I did not directly reflect the enzyme activity of CPT-I. On the other hand, the effects of genistein and genistein with carnitine on the expressions of ACS and CPT-I in muscle were not significant. Our study suggests that genistein with carnitine exerts anti-obesity effects, probably by modulating peroxisome proliferator-activated receptor-associated genes. However, further work is needed to elucidate the possible mechanisms by which genistein and carnitine intervene.

A novel 78-kDa fatty acyl-CoA synthetase (ACS1) of Babesia bovis stimulates memory CD4 + T lymphocyte responses in B. bovis-immune cattle

Antigen-specific CD4 + T lymphocyte responses contribute to protective immunity against Babesia bovis, however the antigens that induce these responses remain largely unknown. A proteomic approach was used to identify novel B. bovis antigens recognized by memory CD4 + T cells from immune cattle. Fractions obtained from merozoites separated by continuous-flow electrophoresis (CFE) that contained proteins ranging from 20 to 83 kDa were previously shown to stimulate memory CD4 + lymphocyte responses in B. bovis-immune cattle. Expression library screening with rabbit antiserum raised against an immunostimulatory CFE fraction identified a clone encoding a predicted 78 kDa protein. BLAST analysis revealed sequence identity of this B. bovis protein with Plasmodium falciparum fatty acyl coenzyme A synthetase (ACS) family members (PfACS1–PfACS11), and the protein was designated B. bovis acyl-CoA synthetase 1 (ACS1). Southern blot analysis indicated that B. bovis ACS1 is encoded by a single gene, although BLAST analysis of the preliminary B. bovis genome sequence identified two additional family members, ACS2 and ACS3. Peripheral blood lymphocytes and CD4 + T cell lines from B. bovis-immune cattle proliferated significantly against recombinant ACS1 protein, consistent with its predicted involvement in protective immunity. However, immune sera from cattle recovered from B. bovis infection did not react with ACS1, indicating that epitopes may be conformationally dependent.

Effect of starvation on hepatic acyl-CoA synthetase, carnitine palmitoyltransferase-I, and acetyl-CoA carboxylase mRNA levels in rats

Objectives This study investigated the effect of starvation on mRNA levels of hepatic acyl coenzyme A synthetase (ACS), carnitine palmitoyltransferase-I (CPT-I), and acetyl coenzyme A carboxylase (ACC) and on serum concentrations of leptin, insulin, and glucose in male Sprague-Dawley rats. Methods Rats were fed an AIN-76 diet for 5 wk and then assigned to a normal group (NG) and a starvation group (SG). The SG was starved for 48 h and the NG was fasted for 12 h before being killed. Serum and hepatic lipids and serum levels of leptin, insulin, and glucose were determined. Expressions of ACS, CPT-1, and ACC mRNA were assessed in liver. Results Serum concentrations of triacylglycerol and high-density lipoprotein cholesterol in the SG were lower than those in the NG. Serum concentrations of low-density lipoprotein cholesterol in the SG were significantly higher than in the NG. Hepatic concentrations of total lipid in the SG were significantly higher than those in the NG, and triacylglycerol concentrations in the SG were significantly lower than those in the NG. Serum concentrations of leptin and glucose in the SG were significantly lower than those in the NG. The ratio of abdominal fat to total body weight in the SG was lower than that in the NG. Hepatic ACS and CPT-I mRNA levels in the SG were significantly higher than those in the NG, but hepatic ACC mRNA levels were lower in the SG than in the NG. Conclusions We demonstrated that starvation increases hepatic levels of ACS and CPT-I and decreases transcription levels of ACC, implicating increases in fatty acid oxidation. This research demonstrates a coordinated regulation of ACS, CPT-I, and ACC mRNA levels and serves to enhance our understanding of the molecular mechanisms underlying fatty acid metabolism during starvation.

A Dictyostelium long chain fatty acyl coenzyme A-synthetase mediates fatty acid retrieval from endosomes

We have identified a subset of Dictyostelium endosomes that carry a long chain fatty acyl coenzyme A-synthetase (LC-FACS 1) on their cytosolic surface. Immunofluorescence studies and observations using GFP-fusion proteins collectively suggest that LC-FACS 1 associates with endosomes a few minutes after their formation, remains bound through the acidic phase of endocytic maturation and dissociates early in the phase where the endosomal content is neutralised prior to exocytosis. Mutants in the fcsA gene, encoding the LC-FACS 1 protein, were constructed by homologous recombination. These cells show a strong defect in the intracellular accumulation of fatty acids, either taken up together with the liquid medium or bound to the surface of particles. Because the mutant cells are otherwise fully competent for macropinocytosis and phagocytosis, we conclude that the LC-FACS 1 protein mediates the retrieval of fatty acids from the lumen of endosomes into the cytoplasm.

Hepatic gene expression in histologically progressive nonalcoholic steatohepatitis

Although the molecular basis for the pathophysiology of nonalcoholic steatohepatitis (NASH) is poorly understood, insulin resistance and mitochondrial dysfunction are physiologic hallmarks of this condition. We sought evidence of a transcriptional or pretranscriptional basis for insulin resistance and mitochondrial dysfunction through measurement of hepatic gene expression (messenger RNA [mRNA]) using high-density synthetic oligonucleotide microarray analysis (Hu6800 GeneChip, Affymetrix, CA). Global hepatic gene expression was determined in snap-frozen liver biopsy specimens from 4 groups: (1) patients with cirrhotic-stage NASH (n = 6), (2) patients with cirrhosis caused by hepatitis C virus (HCV) (n = 6), (3) patients with cirrhosis secondary to primary biliary cirrhosis (PBC) (n = 6), and (4) healthy controls (n = 6). Genes were considered to be expressed differentially in NASH only if there was a greater than 2-fold difference in abundance of mRNA when compared with each of the control groups. Sixteen genes were uniquely differentially expressed (4 overexpressed and 12 underexpressed) in patients with cirrhotic-stage NASH. Genes that were significantly underexpressed included genes important for maintaining mitochondrial function (copper/zinc superoxide dismutase, aldehyde oxidase, and catalase). Glucose 6-phospatase, alcohol dehydrogenase, elongation factor-TU, methylglutaryl coenzyme A (CoA), acyl CoA synthetase, oxoacyl CoA thiolase, and ubiquitin also were underexpressed in NASH. Genes that were overexpressed in NASH included complement component C3 and hepatocyte-derived fibrinogen-related protein, potentially contributing to impaired insulin sensitivity. In conclusion, these studies provide evidence for a transcriptional or pretranscriptional basis for impaired mitochondrial function (attenuated capacity for the dismutation of reactive oxygen species) and diminished insulin sensitivity (increased acute phase reactants) in patients with histologically progressive NASH. Further studies are required to determine the mechanism and the physiologic significance of these findings. (H epatology 2003;38:244-251.)

A nuclear complex containing PPARalpha/RXRalpha is markedly downregulated in the hypertrophied rat left ventricular myocardium with normal systolic function.

The expression of genes encoding fatty acid utilization enzymes is coordinately downregulated during the development of cardiac hypertrophy and failure. However, molecular mechanisms that mediate this downregulation are unknown. Peroxisome proliferator-activated receptor (PPAR) response elements (PPREs) have been identified in promoters of many genes involved in fatty acid utilization, where they function as positive regulatory elements. PPARs bind to PPREs as heterodimers with retinoid X receptors (RXRs). Primary cardiac myocytes from neonatal rats were transfected with a reporter construct driven by the C promoter of rat acyl-coenzyme A synthetase (ACS) gene. Stimulation with phenylephrine, a potent inducer of hypertrophy, markedly downregulated the activity of this promoter. By use of electrophoretic mobility-shift assays (EMSAs) using PPRE in the rat ACS promoter as a probe, we found a sequence-specific protein-DNA complex in the nuclear extract from adult rat left ventricular (LV) myocardium. Supershift experiments revealed that this complex was immunoreactive for PPARalpha and RXRalpha. We compared the activity of this complex in LV nuclear extracts from Dahl salt-sensitive rats (DSs) with hypertension and control age-matched Dahl salt-resistant rats (DRs). Even at the stage of concentric LV hypertrophy with normal systolic function, the activity of the band was markedly diminished in DSs compared with DRs. However, immunoblot analyses showed no difference in LV expression levels of PPARalpha or RXRalpha between DSs and DRs. These findings indicate that a nuclear complex of PPARalpha/RXRalpha is present in adult rat LV and is markedly downregulated in the hypertrophied LV from DS rats, which may account for the loss of transcriptional activation. The downregulation of this complex precedes LV systolic dysfunction and is mediated at the posttranslational levels.

Enhanced expression of hepatic acyl-coenzyme A synthetase and microsomal triglyceride transfer protein messenger RNAs in the obese and hypertriglyceridemic rat with visceral fat accumulation.

The liver plays a central role in lipoprotein metabolism. In particular, very-low density lipoprotein (VLDL) is assembled in the hepatocytes and secreted into the blood circulation. The VLDL is then catabolized to low-density lipoprotein by lipoprotein lipase and hepatic triglyceride lipase. Obese subjects, especially those with visceral fat accumulation, are frequently associated with hyperlipidemia, non-insulin-dependent diabetes mellitus (NIDDM), and hypertension. The mechanism of hyperlipidemia in visceral fat obesity has not yet been elucidated. Otsuka Long-Evans Tokushima Fatty (OLETF) rat is an animal model of NIDDM, characterized by obesity with visceral fat accumulation, hyperlipidemia, and late-onset insulin resistance. To elucidate the mechanism of hyperlipidemia observed in OLETF rats, we focused on the production of VLDL by the liver and investigated hepatic messenger RNA (mRNA) levels of microsomal triglyceride transfer protein (MTP), acyl-coenzyme A synthetase (ACS), and apolipoprotein B (apo B), which play important roles in VLDL synthesis and secretion. In 6-week-old OLETF rats, in which insulin resistance had not been manifested, visceral fat weight was already higher and portal free fatty acid (FFA) and VLDL-triglyceride levels were elevated compared with the control rats. Hepatic ACS activity and mRNA levels, and MTP mRNA levels were also increased in OLETF rats, whereas apo B mRNA levels were similar; these results suggest that the enhanced expression of both ACS and MTP genes associated with visceral fat accumulation before developing insulin resistance may be involved in the pathogenesis of hyperlipidemia in obese animal models with NIDDM.

Effects of gemfibrozil administration on very low density lipoprotein receptor mRNA levels in rabbits

To elucidate the regulation of very low density lipoprotein (VLDL) receptor gene expression, we administered to rabbits for 14 days gemfibrozil, a fibric acid derivative and a lipid lowering drug that is also included among peroxisome proliferators. VLDL receptor mRNA levels were examined by Northern blot analysis. The VLDL receptor mRNA levels in retroperitoneal adipose tissue and in gastrocnemius muscle were increased 6.9-fold and 3.7-fold, respectively, with gemfibrozil treatment, but no marked changes were observed in the heart, the organ in which VLDL receptor is most highly expressed. In the liver, VLDL receptor mRNA was not detected either before or after gemfibrozil administration. Lipoprotein lipase (LPL) and long-chain acyl coenzyme A synthetase (ACS) mRNA levels were also increased in parallel in adipose tissue. The enhanced expression of VLDL receptor mRNA may contribute to the increase of triglyceride-rich lipoprotein catabolism in peripheral tissues such as adipose tissue and muscles.

Induction of the Acyl-Coenzyme A Synthetase Gene by Fibrates and Fatty Acids Is Mediated by a Peroxisome Proliferator Response Element in the C Promoter

The long-chain acyl-coenzyme A synthetase (ACS) gene gives rise to three transcripts containing different first exons preceded by specific regulatory regions A, B, and C. Exon-specific oligonucleotide hybridization indicated that only A-ACS mRNA is expressed in rat liver. Fibrate administration induced liver C-ACS strongly and A-ACS mRNA to a lesser extent. B-ACS mRNA remained undetectable. In primary rat hepatocytes and Fa-32 hepatoma cells C-ACS mRNA increased after treatment with fenofibric acid, alpha-bromopalmitate, tetradecylthioacetic acid, or alpha-linolenic acid. Nuclear run-on experiments indicated that fenofibric acid and alpha-bromopalmitate act at the transcriptional level. Transient transfections showed a 3.4-, 2.3-, and 2.2-fold induction of C-ACS promoter activity after fenofibric acid, alpha-bromopalmitate, and tetradecylthioacetic acid, respectively. Unilateral deletion and site-directed mutagenesis identified a peroxisome proliferator activator receptor (PPAR)-responsive element (PPRE) mediating the responsiveness to fibrates and fatty acids. This ACS PPRE contains three imperfect half sites spaced by 1 and 3 oligonucleotides and binds PPAR.retinoid X receptor heterodimers in gel retardation assays. In conclusion, the regulation of C-ACS mRNA expression by fibrates and fatty acids is mediated by PPAR.retinoid X receptor heterodimers interacting through a PPRE in the C-ACS promoters. PPAR therefore occupies a key position in the transcriptional control of a pivotal enzyme controlling the channeling of fatty acids into various metabolic pathways.

Induction of acyl-coenzyme A synthetase (ACS) gene expression by fibrates is mediated by a PPAR-responsive element in the 1C promoter of the ACS gene

Induction of acyl-coenzyme A synthetase (ACS) gene expression by fibrates is mediated by a PPAR-responsive element in the 1C promoter of the ACS gene

Regulation of eicosanoid synthesis in microvessel endothelium: Glucocorticoids do not affect arachidonyl CoA synthetase activity

The properties of acyl coenzyme A (CoA) synthetase activity were characterized in cultured rabbit coronary microvessel endothelial cells. We report here that microvessel endothelial cells contain two long-chain acyl CoA synthetases. One shows activity with a variety of fatty acids, similar to long-chain non-selective fatty acyl CoA synthetases described previously. The other activity was selective for arachidonic acid and other structurally related substrates. Both activities required ATP, Mg 2+ and CoA for optimal activity. The arachidonyl CoA and the non-selective acyl CoA synthetases showed different thermolabilities. Arachidonyl CoA formation was inhibited by greater than 50% after 1 min at 45°C, whereas a 15 min heating treatment was necessary to produce the same relative inhibition of oleoyl CoA synthesis. Glucocorticoid pretreatment (10 −7 M dexamethasone) of the RCME cells did not affect the apparent K m or V max , nor the fatty acid selectivity for either acyl CoA synthetase. Therefore, although fatty acyl CoA synthetases may be involved in limiting eicosanoid formation, these activities do not appear to be glucocorticoid-responsive.

Inhibition of Neutral Lipase from Castor Bean Lipid Bodies by Coenzyme A (CoA) and Oleoyl-CoA

The neutral lipase (EC 3.1.1.3) in lipid body membranes isolated from the endosperm of 4 day old castor (Ricinus communis L.) seedlings catalyzes the hydrolysis of [(14)C]trioleoylglycerol, releasing [(14)C]oleic acid for up to 4 hours. However, the addition of Mg-ATP and coenzyme A (CoA), which are present in the cytoplasm of plant cells, caused a progressive inhibition of the neutral lipase such that after 15 minutes, release of [(14)C]oleic acid was almost undetectable. A fatty acyl CoA synthetase was found in the lipid body membrane which converts [(14)C]oleic acid produced from the lipase reaction to [(14)C]oleoyl-CoA under these conditions. The concentration of free oleoyl-CoA in the reaction mixture when the lipase was inhibited by 50% was calculated to be about 21 micromolar. It was found that a mixture of exogenously added oleoyl-CoA and CoA was most effective in causing lipase inhibition. Little inhibition of lipase was detected in the presence of CoA alone. It is possible that this effect is important In vivo in coordinating lipase activity with fatty acid oxidation.

Sensor for free fatty acids based on acyl coenzyme-a synthetase and acyl coenzyme-a oxidase

The measurement is based on monitoring dissolved oxgyen consumed by the two sequential reactions catalyzed by the enzymes immobilized in photo-cross-linkable poly (vinyl alcohol) resin (PVA-SbQ). A linear correlation was observed between current decrease and 0.3–2.6 mM oleic or palmitic acid. The sensor was stable in use for three days and could be used to monitor the freshness of oily food.