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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A radiolabeled ATP assay was developed for measuring carboxylic acid:CoA ligase activity. The assay was designed to measure the formation of [gamma-33P]pyrophosphate from [gamma-33P]ATP in the course of the reaction. The assay was linear with protein concentration, and rates as low as 1 pmol/min were measurable. Rates determined with this assay were in agreement with rates determined with [14C]carboxylic acids. The assay was used to characterize the substrate specificity of the XL-I, XL-II, and XL-III ligases from bovine liver mitochondria. Forty carboxylic acids were tested for activity. The enzymes differed in their substrate specificities with XL-I and XL-II being the most similar and XL-III having the broadest specificity. This study has uncovered 19 new carboxylic acids that are substrates for these enzymes.
J Biochem Mol Toxicol 1998
PMID:Development of a radiolabeled ATP assay for carboxylic acid:CoA ligases and its use in the characterization of the xenobiotic carboxylic acid:CoA ligases of bovine liver mitochondria. 952 74

In this work, an attempt was made to identify the reasons of impaired long-chain fatty acid utilization that was previously described in volume-overloaded rat hearts. The most significant data are the following: (1) The slowing down of long-chain fatty acid oxidation in severely hypertrophied hearts cannot be related to a feedback inhibition of carnitine palmitoyltransferase I from an excessive stimulation of glucose oxidation since, because of decreased tissue levels of L-carnitine, glucose oxidation also declines in volume-overloaded hearts. (2) While, in control hearts, the estimated intracellular concentrations of free carnitine are in the range of the respective Km of mitochondrial CPT I, a kinetic limitation of this enzyme could occur in hypertrophied hearts due to a 40% decrease in free carnitine. (3) The impaired palmitate oxidation persists upon the isolation of the mitochondria from these hearts even in presence of saturating concentrations of L-carnitine. In contrast, the rates of the conversion of both palmitoyl-CoA and palmitoylcarnitine into acetyl-CoA are unchanged. (4) The kinetic analyses of palmitoyl-CoA synthase and carnitine palmitoyltransferase I reactions do not reveal any differences between the two mitochondrial populations studied. On the other hand, the conversion of palmitate into palmitoylcarnitine proves to be substrate inhibited already at physiological concentrations of exogenous palmitate. The data presented in this work demonstrate that, during the development of severe cardiac hypertrophy, a fragilization of the mitochondrial outer membrane may occur. The functional integrity of this membrane seems to be further deteriorated by increasing concentrations of free fatty acids which gives rise to an impaired cooperation between palmitoyl-CoA synthase and carnitine palmitoyltransferase I. In intact myocardium, the utilization of the in situ generated palmitoyl-CoA can be further slowed down by decreased intracellular concentrations of free carnitine.
Mol Cell Biochem 1998 Mar
PMID:Palmitate oxidation by the mitochondria from volume-overloaded rat hearts. 954 38

Ethylene can be produced by a variety of developmental and environmental factors such as ripening, the plant hormone auxin, and mechanical wounding via a biosynthetic pathway including AdoMet synthase, ACC synthase, and ACC oxidase steps. ACC synthase and ACC oxidase are known to be encoded by multigene families, and are believed to be differentially expressed in response to various stimuli. In mung bean, ACC synthase is encoded by 7 genes, ACS1, ACS2 ACS3, ACS4, ACS5, ACS6, and ACS7, and ACC oxidase by 2 genes, ACO1 and ACO2. In this study, was have investigated differential accumulation of transcripts for ACC synthase and ACC oxidase homologs in etiolated mung bean hypocotyls under various conditions by the semiquantitative RT-PCR method. Primers which can specifically bind and amplify each cDNAs of ACS1, ACS2, ACS3, ACS4, ACS5, ACS6, ACS7, and ACO1, and ACO2 were designed and used to monitor the responses to various stimuli. Transcripts of ACO1 and ACO2 were accumulated constitutively in the hypocotyl segments even without andy treatment. After cold treatment on intact plant, transcripts of ACS5, ACS6, and ACS7 were accumulated in the hypocotyl segments. We also found the excision of hypocotyl segments and incubation in a buffer solution, a typical way of chemical treatments to hypocotyl segments, lowered the level of ACO2 transcripts with little change of the level of ACO1 transcripts. In response to incubation with IAA (0.1 mM) of excised hypocotyl segments, transcripts of ACS1, ACS6, and ACS7 were accumulated and the level of ACO2 transcripts was increased. Transcripts of ACS1, ACS2, ACS3, ACS5, ACS6 and ACS7 were induced by incubation with OGA (50 micrograms/ml), while the transcripts of ACS4 were accumulated and the level of ACO2 transcripts was increased by incubation with 1 mM LiCl. Our results strongly suggest that all seven ACC synthase genes and two ACC oxidase genes must be active and each gene is differentially regulated by a different subset of the inducing factors.
Mol Cells 1998 Jun 30
PMID:Differential accumulation of transcripts for ACC synthase and ACC oxidase homologs in etiolated mung bean hypocotyls in response to various stimuli. 966 74

Differentially expressed genes generated by cholesterol-loading in the culture medium of aortic smooth muscle cells (SMC) were screened using the DDRT-PCR technique in order to identify the genes that are possibly involved in the pathogenesis of atherosclerosis in the artery. Twenty-eight genes were initially isolated and three differentially expressed cDNAs were finally selected by Northern blot analysis. All three cDNAs were up-regulated (designated CRGSM-1 through -3) by the cholesterol-loading. Upon nucleotide sequencing and homology search in the databases, the first cDNA (CRGSM-1) had a high homology (97%) with the corresponding segment of the acyl-CoA synthetase II gene from rat brain, which participates in fatty acid synthesis. The second one (CRGSM-2) had a high homology (91%) with a part of Mus musculus (mouse) LIM protein 1, and with human skeletal muscle LIM-protein 1 genes (80%) and the third gene (CRGSM-3) had no significant homology match in the database. A full size cDNA isolated from the cDNA library of rat aortic smooth muscle cell using the CRGSM-2 as a probe was identified to have a high homology with muscle LIM protein (MLP). The isolated cDNA contained a segment of DNA that encodes for a zinc-finger motif and two LIM domains. Proteins bearing the LIM domain, defined as a unique double zinc-finger structure associated with a subclass of proteins involved in the determination of cell identity, cell differentiation and control of cell growth, have previously been suggested to play an important role in the pathogenesis of atherosclerosis by others.
Mol Cells 1998 Dec 31
PMID:Differentially expressed genes in cultured aortic smooth muscle cells by cholesterol-loading. 989 16

The petals of daylily (Hemerocallis hybrid) have a genetically based program that leads to senescence and cell death ca. 24 h after the flower opens. In order to determine the components of this program, six cDNAs, whose levels increase during petal senescence, were isolated and sequenced and designated DSA3, 4, 5, 6, 12 and 15. All six DSAs are members of gene families and all but DSA5 and DSA6 have one to three other very similar genes. GenBank database homology searches indicate that DSA3 is most similar at the amino acid level to an in-chain fatty acid hydroxylase which is bound to cytochrome P450, DSA4 may be an aspartic proteinase, DSA5 is as yet unidentified, DSA6 is a putative S1-type nuclease, DSA12 is very similar to a cytochrome P450-containing allene oxide synthase, and DSA15 may be a fatty acid elongase. Except for DSA12, the genes are expressed at low levels in daylily roots. Levels of the DSA mRNAs in leaves are less than 4% of the maximum detected in petals, and there are no clear differences between younger and older leaves. With the exception of DSA4, accumulation of the DSA mRNAs is increased 3.2 to 43 times by a concentration of abscisic acid that causes premature senescence of the petals. The relationship of the putative DSA gene products to senescence and cell death of daylily petals is discussed.
Plant Mol Biol 1999 May
PMID:Identification of senescence-associated genes from daylily petals. 1041 3

Plasmodium is unable to carry out de novo fatty acid synthesis and has to obtain these compounds from their host for subsequent activation by thioesterification with coenzyme A. This activity is catalyzed by a fatty acyl-CoA synthetase enzyme (EC 6.2.1.3). Here, we describe a novel gene from P. falciparum whose recombinant purified product from baculovirus-transfected insect cell line had the enzymatic activity of a long-chain fatty acyl-CoA synthetase. It was named pf acs1, since it belongs to a multi-member gene family as revealed by the sequence of several clones and a multi-band pattern in Southern blots. The sequence specifies a product of 820 amino acid residues. It was transcribed and expressed in infected erythrocytes having an apparent molecular mass of 100 kDa. Immuno-labeling of infected erythrocytes with a specific antibody against the carboxy-terminal part of the PfACS1 localized the product early after the erythrocyte invasion in vesicle-like structures budding off the parasitoforous membrane toward the red cell cytoplasm. Its unique carboxy- terminal structure of 70 extra amino acid residues, longer than any other reported acyl-CoA synthetase, is probably related to its localization in the cytoplasm of the host erythrocyte. The phylogenetic relationship among other AMP-forming enzymes, placed PfACS1 closer to Saccharomyces cerevisiae, sharing significant amino acid identities, especially in the conserved signature motif that modulates fatty acid substrate specificity and ATP/AMP-binding domains. Taking into account the importance of this enzymatic activity for the parasite, its extra-cellular location inside the infected erythrocyte, and the divergence with respect to the homologous human enzymes, it may be an important protein as a potential target candidate for chemotherapeutic antimalaria drugs.
J Mol Biol 1999 Aug 06
PMID:The cloning and expression of Pfacs1, a Plasmodium falciparum fatty acyl coenzyme A synthetase-1 targeted to the host erythrocyte cytoplasm. 1043 6

Activation of fatty acids, catalyzed by acyl-coenzyme A (acyl-CoA) synthetases, is required for their subsequent metabolism. Peroxisomes and microsomes contain very-long-chain acyl-CoA synthetases (VLCSs) capable of activating fatty acids with a chain length of 22 or more carbons. Decreased peroxisomal VLCS activity is, in part, responsible for the biochemical pathology in X-linked adrenoleukodystrophy (X-ALD), illustrating the importance of VLCSs in cellular fatty acid homeostasis. We previously cloned two human genes encoding proteins homologous to rat peroxisomal VLCS; one (hVLCS) is the human ortholog to the rat VLCS gene and another (hVLCS-H1) encodes a related heart-specific protein. Here, we report the cloning of a third gene (hVLCS-H2) and characterization of its protein product. The hVLCS-H2 gene is located on human chromosome 19 and encodes a 690-amino-acid protein. The amino acid sequence of hVLCS-H2 is 44-45% identical and 67-69% similar to those of both hVLCS and hVLCS-H1. COS-1 cells transiently overexpressing hVLCS-H2 activated the very-long-chain fatty acid lignocerate (C24:0) at a rate >1.5-fold higher than that of nontransfected cells (P < 0.002). The hVLCS-H2-dependent activation of long- and branched-chain fatty acids following transient transfection was less striking. However, hVLCS-H2-dependent acyl-CoA synthetase activity with long- and very-long-chain fatty acid substrates was detected in COS-1 cells stably expressing hVLCS-H2. For all substrates tested (C18:0, C20:0, C24:0, C26:0), the hVLCS-H2 catalyzed activity was significantly increased (P < 0.01 to P < 0.0001). By both Northern analysis and reverse transcription polymerase chain reaction, hVLCS-H2 is expressed primarily in liver. Indirect immunofluorescence of COS-1 cells or human hepatoma-derived HepG2 cells expressing epitope-tagged hVLCS-H2 revealed that the protein was associated with the endoplasmic reticulum but not with peroxisomes. Thus, the primary role of hVLCS-H2 is likely to be in fatty acid elongation or complex lipid synthesis rather than in degradation.
Mol Genet Metab 1999 Sep
PMID:Human liver-specific very-long-chain acyl-coenzyme A synthetase: cDNA cloning and characterization of a second enzymatically active protein. 1047 80

The XL-I form of xenobiotic-metabolizing medium-chain fatty acid:CoA ligase was previously purified to apparent homogeneity from bovine liver mitochondria, and the amino acid sequence of a short segment of the enzyme was determined. This sequence was used to develop a probe for screening a bovine cDNA library from which a 1.6 kb cDNA was isolated. This cDNA was sequenced and found to contain the code for the known amino acid sequence. The complete open reading frame was not present in this cDNA, but it was estimated to code for approximately 75% of the XL-I sequence. The XL-III ligase was purified to apparent homogeneity from bovine liver mitochondria. The enzyme eluted from a gel filtration column as a single peak with an apparent molecular weight of ca. 55,000. It ran as a single band on SDS-polyacrylamide gel electrophoresis (SDS-PAGE) with an apparent molecular weight of 62 kDa. N-Terminal sequence analysis of the enzyme gave no sequence, which indicates a blocked N-terminus. The enzyme was chemically cleaved using CNBr. The resulting peptides were separated by SDS-PAGE. The cleavage pattern revealed two large peptides of ca. 21 and 25 kDa, plus several smaller peptides including a prominent 6 kDa peptide. The N-terminus of the 6, 21, and 25 kDa peptides was sequenced and the 21 and 25 kDa sequences were identical indicating incomplete cleavage. The sequences were used to design probes for screening a bovine liver cDNA library. This resulted in the isolation of a 2,065 bp cDNA. This cDNA was sequenced and found to contain the initiation and termination codons, as well as the requisite amino acid sequences. The open reading frame coded for a 64,922 Da protein. The sequence of XL-III cDNA was markedly different from that of XL-I, indicating the genetic uniqueness of the two ligases. They are, however, 64% homologous, which suggests a common evolutionary origin.
J Biochem Mol Toxicol 2000
PMID:Isolation and sequencing of cDNAs for the XL-I and XL-III forms of bovine liver xenobiotic-metabolizing medium-chain fatty acid:CoA ligase. 1056 Oct 77

The deacylation-reacylation cycle is an important mechanism responsible for the introduction of polyunsaturated fatty acids into neural membrane glycerophospholipids. It involves four enzymes, namely acyl-CoA synthetase, acyl-CoA hydrolase, acyl-CoA: lysophospholipid acyltransferase, and phospholipase A2. All of these enzymes have been purified and characterized from brain tissue. Under normal conditions, the stimulation of neural membrane receptors by neurotransmitters and growth factors results in the release of arachidonic acid from neural membrane glycerophospholipids. The released arachidonic acid acts as a second messenger itself. It can be further metabolized to eicosanoids, a group of second messengers involved in a variety of neurochemical functions. A lysophospholipid, the second product of reactions catalyzed by phospholipase A2, is rapidly acylated with acyl-CoA, resulting in the maintenance of the normal and essential neural membrane glycerophospholipid composition. However, under pathological situations (ischemia), the overstimulation of phospholipase A2 results in a rapid generation and accumulation of free fatty acids including arachidonic acid, eicosanoids, and lipid peroxides. This results in neural inflammation, oxidative stress, and neurodegeneration. In neural membranes, the deacylation-reacylation cycle maintains a balance between free and esterified fatty acids, resulting in low levels of arachidonic acid and lysophospholipids. This is necessary for not only normal membrane integrity and function, but also for the optimal activity of the membrane-bound enzymes, receptors, and ion channels involved in normal signal-transduction processes.
J Mol Neurosci 2000 Jun
PMID:Deacylation and reacylation of neural membrane glycerophospholipids. 1098 88

A temperature-sensitive Saccharomyces cerevisiae mutant harboring a lesion in the ERG26 gene has been isolated. ERG26 encodes 4alpha-carboxysterol-C3 dehydrogenase, one of three enzymatic activities required for the conversion of 4,4-dimethylzymosterol to zymosterol. Gas chromatography/mass spectrometry analyses of sterols in this mutant, designated erg26-1, revealed the aberrant accumulation of a 4-methyl-4-carboxy zymosterol intermediate, as well as a novel 4-carboxysterol. Neutral lipid radiolabeling studies showed that erg26-1 cells also harbored defects in the rate of biosynthesis and steady-state levels of mono-, di-, and triglycerides. Phospholipid radiolabeling studies showed defects in the rate of biosynthesis of both phosphatidic acid and phosphatidylinositol. Biochemical studies revealed that microsomes isolated from erg26-1 cells contained greatly reduced 4alpha-carboxysterol-C3 dehydrogenase activity when compared with microsomes from wild type cells. Previous studies have shown that loss of function mutations in either of the fatty acid elongase genes SUR4/ELO3 or FEN1/GNS1/ELO2 can "bypass" the essentiality of certain ERG genes (Ladeveze, V., Marcireau, C., Delourme, D., and Karst, F. (1993) Lipids 28, 907-912; Silve, S., Leplatois, P., Josse, A., Dupuy, P. H., Lanau, C., Kaghad, M., Dhers, C., Picard, C., Rahier, A., Taton, M., Le Fur, G., Caput, D., Ferrara, P., and Loison, G. (1996) Mol. Cell. Biol. 16, 2719-2727). Studies presented here have shown that this sphingolipid-dependent "bypass" mechanism did not suppress the essential requirement for zymosterol biosynthesis. However, studies aimed at understanding the underlying physiology behind the temperature-sensitive growth defect of erg26-1 cells showed that the addition of several antifungal compounds to the growth media of erg26-1 cells could suppress the temperature-sensitive growth defect. Fluorescence microscopic analysis showed that GFP-Erg26p and GFP-Erg27p fusion proteins were localized to the endoplasmic reticulum. Two-hybrid analysis indicated that Erg25p, Erg26p, and Erg27p, which are required for the biosynthesis of zymosterol, form a complex within the cell.
 ...
PMID:The effect of the erg26-1 mutation on the regulation of lipid metabolism in Saccharomyces cerevisiae. 1127 45


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