The Experts below are selected from a list of 204 Experts worldwide ranked by ideXlab platform
Deborah M Muoio - One of the best experts on this subject based on the ideXlab platform.
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the acetyl group buffering action of Carnitine Acetyltransferase offsets macronutrient induced lysine acetylation of mitochondrial proteins
Cell Reports, 2016Co-Authors: Michael N Davies, Lilja Kjalarsdottir, Will J Thompson, Laura G Dubois, Robert Stevens, Olga Ilkayeva, Julia M Brosnan, Timothy P Rolph, Paul A Grimsrud, Deborah M MuoioAbstract:Lysine acetylation (AcK), a posttranslational modification wherein a two-carbon acetyl group binds covalently to a lysine residue, occurs prominently on mitochondrial proteins and has been linked to metabolic dysfunction. An emergent theory suggests mitochondrial AcK occurs via mass action rather than targeted catalysis. To test this hypothesis, we performed mass spectrometry-based acetylproteomic analyses of quadriceps muscles from mice with skeletal muscle-specific deficiency of Carnitine Acetyltransferase (CrAT), an enzyme that buffers the mitochondrial acetyl-CoA pool by converting short-chain acyl-CoAs to their membrane permeant acylCarnitine counterparts. CrAT deficiency increased tissue acetyl-CoA levels and susceptibility to diet-induced AcK of broad-ranging mitochondrial proteins, coincident with diminished whole body glucose control. Sub-compartment acetylproteome analyses of muscles from obese mice and humans showed remarkable overrepresentation of mitochondrial matrix proteins. These findings reveal roles for CrAT and L-Carnitine in modulating the muscle acetylproteome and provide strong experimental evidence favoring the nonenzymatic carbon pressure model of mitochondrial AcK.
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obesity and lipid stress inhibit Carnitine Acetyltransferase activity
Journal of Lipid Research, 2014Co-Authors: Sarah E Seiler, Olga Ilkayeva, Ola J Martin, Robert C Noland, Dorothy H Slentz, Karen L Debalsi, Christopher B Newgard, Timothy R Koves, Deborah M MuoioAbstract:Carnitine Acetyltransferase (CrAT) is a mitochondrial matrix enzyme that catalyzes the interconversion of acetyl-CoA and acetylCarnitine. Emerging evidence suggests that this enzyme functions as a positive regulator of total body glucose tolerance and muscle activity of pyruvate dehydrogenase (PDH), a mitochondrial enzyme complex that promotes glucose oxidation and is feedback inhibited by acetyl-CoA. Here, we used tandem mass spectrometry-based metabolic profiling to identify a negative relationship between CrAT activity and muscle content of lipid intermediates. CrAT specific activity was diminished in muscles from obese and diabetic rodents despite increased protein abundance. This reduction in enzyme activity was accompanied by muscle accumulation of long-chain acylCarnitines (LCACs) and acyl-CoAs and a decline in the acetylCarnitine/acetyl-CoA ratio. In vitro assays demonstrated that palmitoyl-CoA acts as a direct mixed-model inhibitor of CrAT. Similarly, in primary human myocytes grown in culture, nutritional and genetic manipulations that promoted mitochondrial influx of fatty acids resulted in accumulation of LCACs but a pronounced decrease of CrAT-derived short-chain acylCarnitines. These results suggest that lipid-induced antagonism of CrAT might contribute to decreased PDH activity and glucose disposal in the context of obesity and diabetes.
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muscle specific deletion of Carnitine Acetyltransferase compromises glucose tolerance and metabolic flexibility
Cell Metabolism, 2012Co-Authors: Deborah M Muoio, Michael N Davies, Robert Stevens, Olga Ilkayeva, Sarah E Seiler, Robert C Noland, Karen L Debalsi, Jean Paul Kovalik, Indu Kheterpal, Jingying ZhangAbstract:The concept of "metabolic inflexibility" was first introduced to describe the failure of insulin-resistant human subjects to appropriately adjust mitochondrial fuel selection in response to nutritional cues. This phenomenon has since gained increasing recognition as a core component of the metabolic syndrome, but the underlying mechanisms have remained elusive. Here, we identify an essential role for the mitochondrial matrix enzyme, Carnitine Acetyltransferase (CrAT), in regulating substrate switching and glucose tolerance. By converting acetyl-CoA to its membrane permeant acetylCarnitine ester, CrAT regulates mitochondrial and intracellular carbon trafficking. Studies in muscle-specific Crat knockout mice, primary human skeletal myocytes, and human subjects undergoing L-Carnitine supplementation support a model wherein CrAT combats nutrient stress, promotes metabolic flexibility, and enhances insulin action by permitting mitochondrial efflux of excess acetyl moieties that otherwise inhibit key regulatory enzymes such as pyruvate dehydrogenase. These findings offer therapeutically relevant insights into the molecular basis of metabolic inflexibility.
Atsuo Tanaka - One of the best experts on this subject based on the ideXlab platform.
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sorting of peroxisomal and mitochondrial Carnitine Acetyltransferase isozymes in the diploid yeast candida tropicalis
Cell Biochemistry and Biophysics, 2000Co-Authors: Atsuo Tanaka, Mitsuyoshi UedaAbstract:Peroxisomal and mitochondrial Carnitine Acetyltransferase (CAT; EC 2.3.1.7) isozymes are synthesized from the first and second ATG codons of the open reading frame of one gene, Candida tropicalis CAT. Primer extension analysis and RNase protection assay revealed that the peroxisomal CAT, initiating at the second AUG codon of the transcripts, was synthesized by a translational readthrough of the first AUG codon of the open reading frame. When C. tropicalis CAT was introduced into the other yeast, Saccharomyces cerevisiae, 5′ ends of transcripts were similar to those observed in C. tropicalis. Peroxisomal and mitochondrial CAT isozymes were strongly suggested to occur by the alternative initiation of translation, chiefly dependent on the structure or sequence context of the region from the 5′ end to the second AUG codon; their transcripts harbored sufficient information to bring about alternative initiation of translation in both yeasts. Sorting of peroxisomal and mitochondrial CAT isozymes to their own compartment was carried out by their own targeting sequences, but, before transportation to their destination, their biosyntheses were regulated by alternative initiation of translation.
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individual expression of candida tropicalis peroxisomal and mitochondrial Carnitine Acetyltransferase encoding genes and subcellular localization of the products in saccharomyces cerevisiae
Journal of Biochemistry, 1996Co-Authors: Hiroyuki Kawachi, Mitsuyoshi Ueda, Haruyuki Atomi, Noriko Hashimoto, Keiko Kobayashi, Tomoko Yoshida, Naomi Kamasawa, Masako Osumi, Atsuo TanakaAbstract:In an n-alkane assimilating yeast, Candida tropicalis, Carnitine Acetyltransferase (CAT; EC 2.3.1.7) was localized in both peroxisomes and mitochondria. Both CATs were encoded by one gene, CT-CAT, although the initiation sites of translation were suggested to be different. In the present study, the genes corresponding to the supposed C. tropicalis peroxisomal and mitochondrial CATs, which were truncated from the CT-CAT gene, were individually expressed in Saccharomyces cerevisiae, using the C. tropicalis isocitrate lyase promoter (UPR-ICL), which is inducible by oleic acid in concert with proliferation of peroxisomes in S. cerevisiae [Umemura, K., Atomi, H., Kanai, T., Teranishi, Y., Ueda, M., and Tanaka, A. (1995) Appl. Microbiol. Biotechnol. 43, 489-492]. The 71 kDa precursor of mitochondrial CAT, initiating at the first Met, was found to be processed to the mature size (66 kDa) in S. cerevisiae and immunoelectronmicroscopical observation revealed that this enzyme was localized in mitochondria. On the other hand, 68 kDa CAT, initiating at the second Met (residue No. 19), had no cleavable signal and was translocated into peroxisomes and cytosol, but not into mitochondria. The amino-terminal amino acid sequences of individually expressed CATs were identical to those of CATs isolated from alkane-grown C. tropicalis cells, respectively. These results demonstrated that only the 71 kDa protein yielded the 66 kDa protein and that peroxisomal and mitochondrial CATs arose from the difference in the initiation sites of translation.
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presence of Carnitine Acetyltransferase in peroxisomes and in mitochondria of oleic acid grown saccharomyces cerevisiae
Fems Microbiology Letters, 1993Co-Authors: Haruyuki Atomi, Mitsuyoshi Ueda, Junko Suzuki, Yasushi Kamada, Atsuo TanakaAbstract:Activity of Carnitine Acetyltransferase was detected in glucose- and oleic acid-grown Saccharomyces cerevisiae. Oleic acid-grown cells showed a ten-fold higher activity than glucose-grown cells. Subcellular fractionation of oleic acid-grown cells showed that Carnitine Acetyltransferase was present in peroxisomes, mitochondria, and cytosol. The results suggested the plausible presence of an ‘acetylCarnitine shuttle’ in this yeast, as in the case of an n-alkane-assimilating yeast, Candida tropicalis.
Robert Mckenna - One of the best experts on this subject based on the ideXlab platform.
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structural and mutational characterization of l Carnitine binding to human Carnitine Acetyltransferase
Journal of Structural Biology, 2004Co-Authors: Lakshmanan Govindasamy, Wei Lian, Thomas Kukar, Mavis Agbandjemckenna, Brenda Pedersen, Shouguang Jin, Robert MckennaAbstract:We report the crystal structure of a binary complex of human peroxisomal Carnitine Acetyltransferase and the substrate l-Carnitine, refined to a resolution of 1.8 Angstrom with an R(factor) value of 18.9% (R(free)=22.3%). L-Carnitine binds to a preformed pocket in the active site tunnel of Carnitine Acetyltransferase aligned with His(322). The quaternary nitrogen of Carnitine forms a pi-cation interaction with Phe(545), while Arg(497) forms an electrostatic interaction with the negatively charged carboxylate group. An extensive hydrogen bond network also occurs between the carboxylate group and Tyr(431), Thr(444), and a bound water molecule. Site-directed mutagenesis and kinetic characterization reveals that Tyr(431), Thr(444), Arg(497), and Phe(545) are essential for high affinity binding of L-Carnitine.
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Structure of Human Carnitine Acetyltransferase MOLECULAR BASIS FOR FATTY ACYL TRANSFER
The Journal of biological chemistry, 2003Co-Authors: Lakshmanan Govindasamy, Wei Lian, Thomas Kukar, Mavis Agbandje-mckenna, Robert MckennaAbstract:Abstract Carnitine acyltransferases are a family of ubiquitous enzymes that play a pivotal role in cellular energy metabolism. We report here the x-ray structure of human Carnitine Acetyltransferase to a 1.6-A resolution. This structure reveals a monomeric protein of two equally sized α/β domains. Each domain is shown to have a partially similar fold to other known but oligomeric enzymes that are also involved in group-transfer reactions. The unique monomeric arrangement of the two domains constitutes a central narrow active site tunnel, indicating a likely universal feature for all members of the Carnitine acyltransferase family. Superimposition of the substrate complex of a related protein, dihydrolipoyl trans-acetylase, reveals that both substrates localize to the active site tunnel of human Carnitine Acetyltransferase, suggesting the location of the ligand binding sites for Carnitine and coenzyme A. Most significantly, this structure provides critical insights into the molecular basis for fatty acyl chain transfer and a possible common mechanism among a wide range of acyltransferases utilizing a catalytic dyad.
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crystallization and preliminary x ray crystallographic studies on recombinant human Carnitine Acetyltransferase
Acta Crystallographica Section D-biological Crystallography, 2002Co-Authors: Wei Lian, Lakshmanan Govindasamy, Thomas Kukar, Mavis Agbandjemckenna, Robert MckennaAbstract:In this paper, the purification, crystallization and preliminary X-ray crystallographic studies of human Carnitine Acetyltransferase are reported. Recombinant human Carnitine Acetyltransferase crystals were grown by the hanging-drop vapor-diffusion method and belong to the orthorhombic space group P212121, with unit-cell parameters a = 137.65, b = 84.76, c = 57.65 A and one molecule per asymmetric unit. The intensity data were collected from a cryocooled crystal to 1.6 A resolution using a conventional X-ray source.
Randall L Mynatt - One of the best experts on this subject based on the ideXlab platform.
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proximal tubular cell specific ablation of Carnitine Acetyltransferase causes tubular disease and secondary glomerulosclerosis
Diabetes, 2019Co-Authors: Claudia Kruger, Robert C Noland, Jean Paul Kovalik, Trang Tiffany Nguyen, Chelsea Breaux, Alana Guillory, Margaret Mangelli, Kevin T Fridianto, David H Burk, Randall L MynattAbstract:Proximal tubular epithelial cells are highly energy demanding. Their energy need is covered mostly from mitochondrial fatty acid oxidation. Whether derailments in fatty acid metabolism and mitochondrial dysfunction are forerunners of tubular damage has been suggested but is not entirely clear. Here we modeled mitochondrial overload by creating mice lacking the enzyme Carnitine Acetyltransferase (CrAT) in the proximal tubules, thus limiting a primary mechanism to export carbons under conditions of substrate excess. Mice developed tubular disease and, interestingly, secondary glomerulosclerosis. This was accompanied by increased levels of apoptosis regulator and fibrosis markers, increased oxidative stress, and abnormal profiles of acylCarnitines and organic acids suggesting profound impairments in all major forms of nutrient metabolism. When mice with CrAT deletion were fed a high-fat diet, kidney disease was more severe and developed faster. Primary proximal tubular cells isolated from the knockout mice displayed energy deficit and impaired respiration before the onset of pathology, suggesting mitochondrial respiratory abnormalities as a potential underlying mechanism. Our findings support the hypothesis that derailments of mitochondrial energy metabolism may be causative to chronic kidney disease. Our results also suggest that tubular injury may be a primary event followed by secondary glomerulosclerosis, raising the possibility that focusing on normalizing tubular cell mitochondrial function and energy balance could be an important preventative strategy.
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Carnitine Acetyltransferase in agrp neurons is required for the homeostatic adaptation to restricted feeding in male mice
Endocrinology, 2018Co-Authors: Alexander Reichenbach, Randall L Mynatt, Mathieu Mequinion, Jacqueline Bayliss, Sarah Kathleen Haas Lockie, Moyra B Lemus, Romana Stark, Zane B AndrewsAbstract:Abstract Behavioral adaptation to periods of varying food availability is crucial for survival, and agouti-related protein (AgRP) neurons have been associated with entrainment to temporal restricted feeding. We have shown that Carnitine Acetyltransferase (Crat) in AgRP neurons enables metabolic flexibility and appropriate nutrient partitioning. In this study, by restricting food availability to 3 h/d during the light phase, we examined whether Crat is a component of a food-entrainable oscillator (FEO) that helps link behavior to food availability. AgRP Crat knockout (KO) mice consumed less food and regained less body weight but maintained blood glucose levels during the 25-day restricted feeding protocol. Importantly, we observed no difference in meal latency, food anticipatory activity (FAA), or brown adipose tissue temperature during the first 13 days of restricted feeding. However, as the restricted feeding paradigm progressed, we noticed an increased FAA in AgRP Crat KO mice. The delayed increase in FAA, which developed during the last 12 days of restricted feeding, corresponded with elevated plasma levels of corticosterone and nonesterified fatty acids, indicating it resulted from greater energy debt incurred by KO mice over the course of the experiment. These experiments highlight the importance of Crat in AgRP neurons in regulating feeding behavior and body weight gain during restricted feeding but not in synchronizing behavior to food availability. Thus, Crat within AgRP neurons forms a component of the homeostatic response to restricted feeding but is not likely to be a molecular component of FEO.
Abdullah S. Alhomida - One of the best experts on this subject based on the ideXlab platform.
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Investigations of the Effects of Theophylline Administration on Carnitine Acetyltransferase Activity of Rat Heart
Journal of enzyme inhibition, 1997Co-Authors: Abdullah S. AlhomidaAbstract:AbstractThe effects of oral theophylline administration (100 mg/kgb.w./day) on the activity of Carnitine Acetyltransferase (CAT) of rat heart for five-week interval treatment were studied. The result indicated that the body weights of placebo groups were not significantly changed as compared to control groups (P < 0.1), but theophylline treatment caused a significant decrease in the body weights of rat (P < 0.01) as compared to either control or placebo groups throughout the five-week interval treatments. Daily administration of theophylline to rats did not significantly affect heart weights as compared to either control or placebo groups (P < 0.1) for the five week-interval treatments. Our data indicated that the activity of CAT was not significantly changed in placebo groups as compared to control groups (P < 0.1), however, there was a significant increase in the activity of CAT in heart of theophylline-treated groups (P < 0.01) as compared to either control or placebo groups. The increase in the activi...
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inhibition studies of the Carnitine Acetyltransferase from skeletal muscle of the camel camelus dromedarius by sulfhydryl reagents and metal ions
Iubmb Life, 1996Co-Authors: Abdullah S. AlhomidaAbstract:The effect of certain sulfhydryl reagents and metal ions were studied on the Carnitine Acetyltransferase (CAT) activity from the skeletal muscle of the Arabian camel (Camelus dromedarius). DTNB and iodoacetamide caused concentration and time dependent inhibition of CAT activity. The inhibition seen with these sulfhydryl reagents could be protected with prior incubation of the enzyme with acetyl-Co A, suggesting that these reagents might interact with the same site. Among the various metal ions tested, Cu2+, Zn2+ and Hg2+ caused total inhibition at very low concentrations, while, Mn2+, Mo6+ and Co2+ caused between 32-52% inhibition at 10 mM concentrations. Alkali earth divalent metals Mg2+ and Ca2+ caused less than 15% inhibition at this concentration. These metal ions are probably interacting at certain nucleophilic groups in the enzyme thus disrupting its tertiary structure.
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kinetic properties of purified Carnitine Acetyltransferase from the skeletal muscle of arabian camel camelus dromedarius
Biochimie, 1996Co-Authors: Abdullah S. Alhomida, A. S. Duhaiman, Abdulaziz A Aljafari, Nayyar Rabbani, M. A. JunaidAbstract:Abstract The kinetic properties of Carnitine Acetyltransferase from the skeletal muscle of the Arabian camel ( Camelus dromedarius ) were studied. The enzyme showed an optimum pH between 7.2 and 8.2. Reciprocal plots of data obtained by varying one substrate concentration while keeping the other constant revealed lines that converged on the abscissa, indicating that the enzyme possibly follows a random mechanism of catalysis. The K m s for L-Carnitine and acetyl-coenzyme A were 244 and 44 μM respectively, while those for acetyl-DL-carnithine and coenzyme A (Co A) were 307 and 39 μM respectively. The K m for one substrate was found to be independent of the concentration of the second substrate used. Corresponding V max values for L-CA, acetyl-Co A, acetyl-DL-Carnitine and Co A are 98, 98, 102 and 100 μmol min −1 mg −1 protein respectively. The low K m obtained for acetyl-DL-Carnitine suggests an adaptive mechanism in this desert species for enduring prolonged dry spells without food and water.