Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.21 (CPT)
 4,580 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A cDNA encoding full-length carnitine palmitoyltransferase I (CPT I) from rat liver was expressed in Saccharomyces cerevisiae, a system devoid of endogenous CPT activity. The recombinant enzyme was of the expected size (as deduced from immunoblots), membrane-bound, and detergent-labile. It was also potently inhibited by malonyl-CoA, with an I50 value (concentration causing 50% inhibition) of approximately 5 microM, similar to that of the native enzyme in rat liver mitochondria. A truncated variant of the enzyme that lacked the amino-terminal 82 residues encompassing the first hydrophobic domain retained catalytic function but was much less sensitive to malonyl-CoA (I50 > 80 microM). Deletion of the cDNA segment encoding amino acids 31-148 (which includes both first and second hydrophobic stretches) resulted in no detectable product. The data establish unequivocally that a single polypeptide possesses both catalytic and malonyl-CoA binding domains, as well as the other properties previously attributed by us to native CPT I in mammalian mitochondria, and should thus put to rest the controversy surrounding this issue (Kerner, J., Zaluzec, E., Gage, D., and Bieber, L. L. (1994) J. Biol. Chem. 269, 8209-8219). In addition, the results strengthen the view that one site of interaction of malonyl-CoA with the rat liver enzyme involves the NH2-terminal region of the molecule.
 ...
PMID:Expression of a cDNA for rat liver carnitine palmitoyltransferase I in yeast establishes that catalytic activity and malonyl-CoA sensitivity reside in a single polypeptide. 792 64

It has recently been established that rat heart mitochondria contain two isoforms of carnitine palmitoyltransferase I (CPT I), the minor 88-kDa variant being identical to liver CPT I (L-CPT I) and the dominant 82-kDa form resembling the skeletal muscle enzyme (M-CPT I) (Weis, B. C., Esser, V., Foster, D. W., and McGarry, J. D. (1994) J. Biol. Chem. 269, 18712-18715). To quantify the functional contribution of L-CPT I to overall CPT I activity in heart mitochondria a selective inhibitor of the former was needed. The dinitrophenol analog of 2[6-(4-chlorophenoxy)hexyl]oxirane-2-carboxylic acid (etomoxir) (DNP-Et) was found to have this property. When liver and skeletal muscle mitochondria were exposed to DNP-Et in the presence of ATP and CoASH, the DNP-Et-CoA formed completely inhibited liver CPT I while leaving the muscle enzyme unaffected. Similar treatment of heart mitochondria blocked only the L-CPT I component. This had the effect of shifting the apparent Km for carnitine from approximately 200 to approximately 500 microM and the I50 value for malonyl-CoA (the concentration needed to suppress enzyme activity by 50%) from approximately 0.18 to approximately 0.06 microM, i.e. the heart system now behaved exactly the same as that from skeletal muscle. Taking the Km for carnitine of L-CPT I and M-CPT I to be 30 and 500 microM, respectively, it could be calculated that the former contributes approximately 2% to the total CPT I in heart. When the 82-kDa CPT I isoforms of heart and skeletal muscle were labeled with [3H]etomoxir and then exposed to trypsin, the fragmentation patterns obtained were identical and quite distinct from that given by CPT I from liver. We conclude that (i) DNP-Et, unlike other agents of the oxirane carboxylic acid class, has remarkable inhibitory selectivity for L-CPT I over M-CPT I; (ii) the previously puzzling observation that rat heart CPT I displays kinetic characteristics intermediate between those of the enzymes from liver and skeletal muscle is entirely accounted for by the low level expression of L-CPT I in the cardiac myocyte; and (iii) the dominant 82-kDa CPT I isoform in heart is identical to the muscle enzyme. The data reaffirm that, in contrast to CPT II, CPT I exists in at least two isoforms and that both are present in rat heart.
 ...
PMID:Use of a selective inhibitor of liver carnitine palmitoyltransferase I (CPT I) allows quantification of its contribution to total CPT I activity in rat heart. Evidence that the dominant cardiac CPT I isoform is identical to the skeletal muscle enzyme. 792 65

The mechanism of activation of mitochondrial overt carnitine palmitoyltransferase (CPT I) by treatment of hepatocytes with okadaic acid (OA) was investigated. Activation was observed when cells were permeabilized with digitonin, but not when a total membrane fraction was obtained by sonication. Both cell disruption methods preserved the activation of phosphorylase observed in OA-treated hepatocytes. Activation of CPT I was also observed in crude homogenates of OA-treated hepatocytes, but it was lost upon subsequent isolation of mitochondria from such homogenates. In all experiments, any activation observed did not depend on the presence or absence of fluoride ions in the permeabilization/homogenization media. When hepatocytes were permeabilized in the absence of fluoride and further incubated with exogenous phosphatases 1 and 2A, the OA-induced activation of CPT was not reversed, whereas the activation of glycogen phosphorylase in the same cells was rapidly reversed. Treatment of hepatocytes with OA, followed by permeabilization and incubation before assay of CPT I, demonstrated that OA had no short-term effect on the sensitivity of CPT I to malonyl-CoA, although the difference in sensitivity between cells isolated from fed and starved rats was fully preserved. Incubation of isolated mitochondria or purified mitochondrial outer membranes with cyclic AMP-dependent or AMP-activated protein kinases, under phosphorylating conditions, did not affect the activity of CPT I or its sensitivity to malonyl-CoA inhibition. Under the same conditions, the use of [32P]ATP resulted in the labelling of several outer-membrane proteins but, unlike [3H]etomoxir-labelled CPT I, none of them was specifically removed from membrane extracts by a specific polyclonal antibody to the enzyme. We conclude that the increase in overt CPT activity observed in permeabilized hepatocytes is not due to direct phosphorylation of CPT I, but may involve interactions between the mitochondrial outer membrane and other membranous or soluble cytosolic components of the cell.
 ...
PMID:Evidence against direct involvement of phosphorylation in the activation of carnitine palmitoyltransferase by okadaic acid in rat hepatocytes. 801 Sep 50

We sought to explore the emerging concept that malonyl-CoA generation, with concomitant suppression of mitochondrial carnitine palmitoyltransferase I (CPT I), represents an important component of glucose-stimulated insulin secretion (GSIS) by the pancreatic beta-cell (Prentki M, Vischer S, Glennon MC, Regazzi R, Deeney JT, Corkey BE: Malonyl-CoA and long-chain acyl-CoA esters as metabolic coupling factors in nutrient-induced insulin secretion. J Biol Chem 267:5802-5810, 1992). Accordingly, pancreases from fed rats were perfused with basal (3 mM) followed by high (20 mM) glucose in the absence or presence of 2 mM hydroxycitrate (HC), an inhibitor of ATP-citrate (CIT) lyase (the penultimate step in the glucose-->malonyl-CoA conversion). HC profoundly inhibited GSIS, whereas CIT had no effect. Inclusion of 0.5 mM palmitate in the perfusate significantly enhanced GSIS and completely offset the negative effect of HC. In isolated islets, HC stimulated [1-14C]palmitate oxidation in the presence of basal glucose and markedly obtunded the inhibitory effect of high glucose. Directional changes in 14C incorporation into phospholipids were opposite to those of 14CO2 production. At a concentration of 0.2 mM, 2-bromostearate, 2-bromopalmitate and etomoxir (all CPT I inhibitors) potentiated GSIS by the pancreas and inhibited palmitate oxidation in islets. However, at 0.05 mM, etomoxir did not influence insulin secretion but still caused significant suppression of fatty acid oxidation. The results provide more direct evidence for a pivotal role of malonyl-CoA suppression of CPT I, with attendant elevation of the cytosolic long-chain acyl-CoA concentration, in GSIS from the normal pancreatic beta-cell.(ABSTRACT TRUNCATED AT 250 WORDS)
 ...
PMID:More direct evidence for a malonyl-CoA-carnitine palmitoyltransferase I interaction as a key event in pancreatic beta-cell signaling. 801 51

To begin to explore the basis for the tissue-specific expression of mitochondrial carnitine palmitoyltransferase I (CPT I), we focused on three rat tissues (liver, heart, and skeletal muscle) in which the enzyme was known to display very different properties. In Northern blot analysis, a cDNA probe corresponding to liver CPT I readily hybridized to a 4.5-kilobase species of mRNA in liver and heart, but not in skeletal muscle. Using the same probe to screen a neonatal rat heart cDNA library, a full-length clone, surprisingly having 100% sequence identity to the liver CPT I cDNA, was isolated. The paradox was resolved by two additional experiments. First, in Western blots of mitochondrial membranes, an antibody raised against liver CPT I recognized the 88-kDa protein in heart, as well as in liver, but not in skeletal muscle. Second, high specific activity [3H]deschloroetomoxir (a covalent ligand for CPT I) reacted with a single form of CPT I in liver (approximately 88 kDa) and skeletal muscle (approximately 82 kDa), while proteins of both sizes were labeled in the cardiac myocyte. Tritiated ligand binding to the two heart proteins was blocked by excess unlabeled malonyl-CoA. It is concluded that liver and skeletal muscle each contains a single and distinct isoform of CPT I with monomeric size of approximately 88 and 82 kDa, respectively. The heart contains a CPT I protein of approximately 82 kDa in size (probably identical to the skeletal muscle protein) but, importantly, also expresses the liver-type enzyme. The results likely explain why previous studies of heart CPT I yielded an apparent Km for carnitine and I50 value for malonyl-CoA inhibition that were intermediate between those of the liver and skeletal muscle enzymes.
 ...
PMID:Rat heart expresses two forms of mitochondrial carnitine palmitoyltransferase I. The minor component is identical to the liver enzyme. 803 22

We describe two male infants with central nervous system disorders, i.e. infantile spasms in one and athetotic quadriplegia in the other, and with recurrent attacks of high plasma creatine kinase levels induced by viral infections. Although carnitine palmitoyltransferase I (CPT I) activity in biopsied muscle was normal in both cases, that of carnitine palmitoyltransferase II (CPT II) was decreased to 37% and 25% of the control value, respectively. Meanwhile, to determine whether or not and how CPT exists in the central nervous system (CNS), we studied animal brain tissues. CPT activity was demonstrated in almost all regions, especially in the brainstem, cerebellum and spinal cord. Although CPT deficiency can be classified into hepatic (CPT I) and muscular (CPT II) presentations, these data suggest that another symptomatology of CPT II deficiency with CNS involvement (brain type?) might exist.
 ...
PMID:Central nervous system disorders and possible brain type carnitine palmitoyltransferase II deficiency. 804 3

The effects of the ingestion of a meal on the partitioning of hepatic fatty acids between oxidation and esterification were studied in vivo for meal-fed rats. The time course for the reversal of the starved state was extremely rapid and the process was complete within 2 h, in marked contrast with the reversal of the effects of starvation in rats fed ad libitum [A. M. B. Moir and V. A. Zammit (1993) Biochem. J. 289, 49-55]. This rapid reversal occurred in spite of the fact that, in the liver of the meal-fed animals before feeding, a similar degree of partitioning of fatty acids in favour of oxidation was observed as in 24 h-starved rats (previously fed ad libitum). This suggested that the lower degree of ketonaemia observed in meal-fed rats before a meal is not due to the inability of acylcarnitine formation to compete successfully with esterification of fatty acids to the glycerol moiety. Investigation of the possible mechanisms that could contribute towards the rapid switching-off of fatty acid oxidation revealed that this was correlated with a very rapid rise and overshoot in hepatic malonyl-CoA concentration, but not with any change in the activity, or sensitivity to malonyl-CoA, of the mitochondrial overt carnitine palmitoyltransferase (CPT I). The role of these two parameters in the reversal of fasting-induced hepatic fatty acid oxidation was thus the inverse of that observed previously for refed 24 h-starved rats. The rapid increase in [malonyl-CoA] was accompanied by an immediate and complete reversion of the kinetic characteristics (Ka for citrate, expressed/total activity ratio) of acetyl-CoA carboxylase to those found in the post-meal animals, again in contrast with the time course observed in refed 24 h-starved rats [A. M. B. Moir and V. A. Zammit (1990) Biochem. J. 272, 511-517]. The rapidity with which these changes occurred was specific to the partitioning of acyl-CoA; the meal-induced diversion of glycerolipids towards phospholipid synthesis and the acute inhibition of the fractional rate of triacylglycerol secretion occurred with very similar time courses to those observed upon refeeding of 24 h-starved rats. The results confirm the central role played by differences in the dynamics of changes in hepatic malonyl-CoA concentration, and CPT I sensitivity to it, in determining the route through which ingested glucose is converted into hepatic glycogen upon refeeding of starved rats which had previously been meal-fed or fed ad libitum.
 ...
PMID:Rapid switch of hepatic fatty acid metabolism from oxidation to esterification during diurnal feeding of meal-fed rats correlates with changes in the properties of acetyl-CoA carboxylase, but not of carnitine palmitoyltransferase I. 809 87

The effect of thermal acclimation on the activity of carnitine palmitoyltransferase I (CPT I), the rate-limiting enzyme for beta-oxidation of long-chain fatty acids, was determined in oxidative red muscle of striped bass (Morone saxatilis) acclimated at 5 or 25 degrees C. As observed in mammalian tissues, malonyl-CoA potently inhibited CPT I activity of mitochondria. Inhibition by malonyl-CoA required inclusions of both bovine serum albumin (BSA) and palmitoyl-CoA in the reaction media. Because BSA binds long-chain fatty acyl-CoAs, this observation suggests that free fatty acyl-CoAs may disrupt mitochondrial membranes and affect the CPT I protein. Cold acclimation increased citrate synthase activity 1.6-fold and total CPT activity 2-fold in homogenates of red muscle; free carnitine increased 62%, and specific activity of CPT I in mitochondria increased 2-fold. No differences were observed between cold- and warm-acclimated fish in substrate-binding properties of CPT I at an assay temperature of 15 degrees C, as judged by the Michaelis constant (Km) for carnitine (0.11 +/- 0.02 vs. 0.13 +/- 0.02 mM) or inhibition of CPT I, as determined by the half-maximal inhibition concentration (IC50) for malonyl-CoA (0.14 +/- 0.05 vs. 0.09 +/- 0.03 microM). Thermal sensitivity of CPT I (Q10 = 2.91 +/- 0.12 vs. 3.02 +/- 0.20) and preference of CPT I for different long-chain fatty acyl-CoA substrates (16:1-CoA = 16:0-CoA > 18:1-CoA) were not altered by thermal acclimation.(ABSTRACT TRUNCATED AT 250 WORDS)
 ...
PMID:Cold acclimation increases carnitine palmitoyltransferase I activity in oxidative muscle of striped bass. 814 97

The review examines the mechanisms regulating the activities of the two key enzymes determining rates of glucose and fatty acid oxidation, i.e., the pyruvate dehydrogenase (PDH) complex and the carnitine palmitoyltransferase (CPT) system. The review also evaluates the regulatory importance of gene expression in the control of tissue fuel selection within the context of substrate competition between glucose and fatty acids. It identifies a strong indirect input of nutrient-gene interactions in the control of pyruvate oxidation through the regulated provision of pyruvate as a substrate for PDH and as an inhibitor of PDH kinase. Nutrient-gene interactions are also identified in relation to the regulation of CPT I activity by malonyl-CoA (inhibitor) and by the provision of long-chain acyl-CoA (substrate/activator), the latter via the hydrolysis of plasma or tissue triacylglycerol (by lipoprotein lipase and hormone-sensitive lipase, respectively). We discuss how such regulation is reinforced by long-term modulation of PDH kinase-specific activity and CPT I maximal activity. We also explore the role of mechanisms operating at the levels of the PDH complex and the CPT system that act to promote and accelerate a switch in fuel utilization once a committed change in nutrient supply has been established. In particular, we discuss the regulatory influences exerted by altered sensitivities of PDH kinase to inhibition by pyruvate and CPT I to inhibition by malonyl-CoA, respectively.
 ...
PMID:Interactive regulation of the pyruvate dehydrogenase complex and the carnitine palmitoyltransferase system. 829 90

1. The technique of selective labelling of hepatic fatty acids in vivo [Moir and Zammit (1992) Biochem. J. 283, 145-149] has been used to monitor non-invasively the metabolism of fatty acids in the livers of awake unrestrained rats during the starved-to-refed transition. Values for the incorporation of labelled fatty acid into liver and plasma glycerolipids and into exhaled carbon dioxide after injection of labelled lipoprotein and Triton WR 1339 into rats with chronically cannulated jugular veins were obtained for successive 1 h periods from the start of refeeding of 24 h-starved rats. 2. Starvation for 24 h resulted in marked and reciprocal changes in the incorporation of label into glycerolipids and exhaled 14CO2, such that a 4-fold higher value was obtained for the oxidation/esterification ratio in livers of starved rats compared with fed animals. 3. Refeeding of starved rats did not return this ratio to the value observed for fed animals for at least 7 h; during the first 3 h of refeeding the ratio was at least as high as that for starved rats. Between 4 h and 6 h of refeeding the ratio was still approx. 70% of that in starved animals, and 2.5-fold higher than in fed rats. 4. These data support the hypothesis that the capacity of the liver to oxidize fatty acids is maintained at a high level during the initial stages of refeeding [Grantham and Zammit (1986) Biochem. J. 239, 485-488] and that control of the flux of hepatic fatty acids into the oxidative pathway is largely lost from the reaction catalysed by mitochondrial overt carnitine palmitoyltransferase (CPT I) during this phase of recovery from the starved state. 5. Refeeding also resulted in a rapid (< 1 h) increase in hepatic malonyl-CoA concentrations to values intermediate between those in livers of fed and starved animals. The sensitivity of CPT I to malonyl-CoA inhibition in isolated liver mitochondria was only partially reversed even after 5 h of refeeding. 6. Refeeding resulted in an acute 35% inhibition of the fraction of synthesized triacylglycerol that was secreted into the plasma; the maximal effect occurred 2-3 h after the start of refeeding. The inhibition of the fractional secretion rate was fully reversed after 5 h of refeeding. 7. The amount of 14C label that was incorporated into phospholipids as a fraction of total glycerolipid synthesis was doubled within 2 h of the start of refeeding.(ABSTRACT TRUNCATED AT 400 WORDS)
 ...
PMID:Monitoring of changes in hepatic fatty acid and glycerolipid metabolism during the starved-to-fed transition in vivo. Studies on awake, unrestrained rats. 842 71


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>