Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.3.1.21 (CPT)
4,580 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The degree of inhibition of CPT I (carnitine palmitoyltransferase, EC 2.3.1.21) in isolated rat liver mitochondria by malonyl-CoA was studied by measuring the activity of the enzyme over a short period (15s) after exposure of the mitochondria to malonyl-CoA for different lengths of time. Inhibition of CPT I by malonyl-CoA was markedly time-dependent, and the increase occurred at the same rate in the presence or absence of palmitoyl-CoA (80 microM), and in the presence of carnitine, such that the time-course of acylcarnitine formation deviated markedly from linearity when CPT I activity was measured in the presence of malonyl-CoA over several minutes. The initial rate of increase in degree of inhibition with time was independent of malonyl-CoA concentration. CPT I in mitochondria from 48 h-starved rats had a lower degree of inhibition by malonyl-CoA at zero time, but was equally capable of being sensitized to malonyl-CoA, as judged by an initial rate of increase of inhibition identical with that of the enzyme in mitochondria from fed rats. Double-reciprocal plots for the degree of inhibition produced by different malonyl-CoA concentrations at zero time for the enzyme in mitochondria from fed or starved animals indicated that the enzyme in the latter mitochondria was predominantly in a state with low affinity for malonyl-CoA (concentration required to give 50% inhibition, I0.5 congruent to 10 microM), whereas that in mitochondria from fed rats displayed two distinct sets of affinities: low (congruent to 10 microM) and high (less than 0.3 microM). Plots for mitochondria after incubation for 0.5 or 1 min with malonyl-CoA indicated that the increased sensitivity observed with time was due to a gradual increase in the high-affinity state in both types of mitochondria. These results suggest that the sensitivity of CPT I in rat liver mitochondria in vitro had two components: (i) an instantaneous sensitivity inherent to the enzyme which depends on the nutritional state of the animal from which the mitochondria are isolated, and (ii) a slow, malonyl-CoA-induced, time-dependent increase in sensitivity. It is suggested that the rate of malonyl-CoA-induced sensitization of the enzyme to malonyl-CoA inhibition is limited by a slow first-order process, which occurs after the primary event of interaction of malonyl-CoA with the mitochondria.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Time-dependence of inhibition of carnitine palmitoyltransferase I by malonyl-CoA in mitochondria isolated from livers of fed or starved rats. Evidence for transition of the enzyme between states of low and high affinity for malonyl-CoA. 671 21

The kinetics of carnitine palmitoyltransferase I (CPT I; EC 2.3.1.21) were examined in mitochondria from rat liver, heart and skeletal muscle as a function of pH over the range 6.8-7.6. In all three tissues raising the pH resulted in a fall in the Km for carnitine, no change in the Km for palmitoyl-CoA or Octanoyl-CoA, and a marked decrease in the inhibitory potency of malonyl-CoA. Studies with skeletal-muscle mitochondria established that increasing pH was accompanied by an increase in the Kd of the malonyl-CoA binding site for this ligand, coupled with a decrease in the Kd for fatty acyl-CoA species to compete for malonyl-CoA binding. Three principal conclusions are drawn. (1) The pH-induced shift in malonyl-CoA sensitivity of CPT I is not a phenomenon restricted to liver mitochondria. (2) At any given pH within the range tested, the ability of malonyl-CoA (and closely related compounds) to inhibit enzyme activity is governed by the efficiency of their binding to the malonyl-CoA site. (3) The competitive interaction between fatty acyl-CoA substrates and malonyl-CoA as regards CPT I activity is exerted at the malonyl-CoA binding site. Finally, the possibility is strengthened that the malonyl-CoA binding site is distinct from the active site of CPT I.
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PMID:Effects of pH on the interaction of substrates and malonyl-CoA with mitochondrial carnitine palmitoyltransferase I. 674 35

The effect of progressive, low-intensity endurance training on regulatory enzyme activities in slow-twitch (ST) and fast-twitch (FT) muscle fibres was studied in 32 rats. Of those rats 16 were trained on a treadmill at a running speed of 10 m.min-1 5 days a week over an 8-week period. Running time was progressively increased from 15 min to 2 h.day-1. Of the rats 4 trained and 4 sedentary rats were also subjected to acute exhausting exercise. Enzyme activities of phosphofructokinase 1 (PFK1) from glycolysis, alpha-ketoglutarate dehydrogenase (alpha-KGDH) from the Krebs cycle and carnitine palmitoyltransferase (CPT I and II) from fatty acid metabolism in soleus, tibialis anterior and gastrocnemius muscles were measured in trained and sedentary rats. Enzyme activities of individual ST and FT fibres were measured from the freeze-dried gastrocnemius muscle of 8 trained and 8 sedentary rats. In the sedentary rats the activity of PFK1 in tibialis anterior and soleus muscles was 141% and 41% of the activity in gastrocnemius muscle, respectively. The activity of alpha-KGDH in tibialis anterior and soleus muscles was 164% and 278% of the activity in gastrocnemius muscle, respectively. The activity of CPT I in tibialis anterior and gastrocnemius muscles were at the same level, but in soleus muscle the activity was 127% of that in mixed muscle. Endurance training increased enzyme activities of alpha-KGDH and CPT I significantly (P < 0.05) in gastrocnemius muscle but not in soleus or tibialis anterior muscle.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Alteration of regulatory enzyme activities in fast-twitch and slow-twitch muscles and muscle fibres in low-intensity endurance-trained rats. 764 37

The expression pattern of mitochondrial carnitine palmitoyltransferase (CPT) enzymes was examined in the developing rat heart. Whereas the specific activity of CPT II increased approximately 3-fold during the first month of life, the profile for CPT I, which is composed of both liver (L) and muscle (M) isoforms, was more complex. Exposure of mitochondria to [3H]etomoxir (a covalent ligand for CPT I), followed by fluorographic analysis of the membrane proteins, established that while in the adult heart L-CPT I represents a very minor constituent, its contribution is much greater in the newborn animal. Use of the related inhibitor, 2-[6-(2,4-dinitrophenoxy)hexyl]oxirane-2-carboxylic acid (specific for L-CPT I), allowed the activities of the two CPT I variants to be quantified separately. The results showed that in the neonatal heart, L-CPT I contributes approximately 25% to total CPT I activity (in Vmax terms), the value falling during growth of the pups (with concomitant increasing expression of the M isoform) to its adult level of 2-3%. Because the myocardial carnitine content is very low at birth and rises dramatically over the next several weeks, it can be estimated that L-CPT I (Km for carnitine of only 30 microM compared with a value of 500 microM for M-CPT I) is responsible for some 60% of total cardiac fatty acid oxidation in the newborn rat; the value falls to approximately 4% in adult animals. Should these findings have a parallel in humans, they could have important implications for understanding the pathophysiological consequences of inherited L-CPT I deficiency syndromes.
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PMID:Mitochondrial carnitine palmitoyltransferase I isoform switching in the developing rat heart. 772 4

This study was conducted to determine if the activity of 2,4-dienoyl-CoA reductase limits the rate of cardiac beta-oxidation of highly unsaturated fatty acids. Although growth hormone treatment of hypophysectomized rats caused a 3-fold increase in the activity of 2,4-dienoyl-CoA reductase, beta-oxidation of docosahexaenoate in cardiomyocytes was not stimulated by this treatment. Since cardiomyocytes oxidized oleic acid more rapidly than docosahexaenoic acid, the utilization of energy did not limit beta-oxidation. Respiration measurements with coupled rat heart mitochondria revealed that the rates of beta-oxidation with palmitoyl-CoA and palmitoylcarnitine as substrates were virtually identical but were 3- to 4-fold higher than the rates obtained with either docosahexaenoyl-CoA or docosahexaenoylcarnitine. Although the activity of carnitine palmitoyltransferase I (CPT I) was 5 times higher with palmitoyl-CoA as substrate than with docosahexaenoyl-CoA, this reaction is only one of several that may limit the beta-oxidation of docosahexaenoic acid. Surprisingly, an incremental inhibition of CPT I resulted in a parallel inhibition of respiration supported by either palmitoyl-CoA or docosahexaenoyl-CoA. This observation agrees with the notion that CPT I may also be a regulatory enzyme in cardiac fatty acid oxidation. It is concluded that the reduction of double bonds by 2,4-dienoyl-CoA reductase does not restrict the cardiac beta-oxidation of highly unsaturated fatty acid, like docosahexaenoic acid.
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PMID:On the rate-limiting step in the beta-oxidation of polyunsaturated fatty acids in the heart. 773 39

The requirement for a normal insulin response in mediating the starved-to-refed transition, with respect to the partitioning of hepatic fatty acids between beta-oxidation and esterification to glycerol, was studied. Diabetic rats were starved for 24 h and refed ad libitum for various periods of time. There was no increase in plasma insulin in response to the meal. However, the fatty acid oxidation:esterification ratio was very rapidly decreased from the starved to the fed value, most of the transition being achieved within the first hour of refeeding. There was a 2 h lag in the response of hepatic malonyl-CoA concentration, such that this rapid switch from oxidation to esterification could not be explained on the basis of changes in the absolute concentration of this inhibitor of carnitine palmitoyltransferase I (CPT I). Hepatic pyruvate and lactate concentrations both increased by several-fold upon refeeding and peaked after 1 h and 3 h, respectively. The hepatic lactate:pyruvate ratio increased 3.2-fold during the first 3 h of refeeding, suggesting that the cytosolic NAD(+)-NADH couple became much more highly reduced during the lag-period between the onset of inhibition of flux of fatty acids towards oxidation and the rise in malonyl-CoA concentration. This may be indicative of a lowering of intracellular pH, which would amplify greatly the sensitivity of CPT I to the inhibitor. In view of the very rapid and high food intake by these diabetic rats, the possibility is also considered that portal concentrations of amino acids and other metabolites could give rise to an increase in liver cell-volume that would inhibit CPT I acutely by an as yet unknown mechanism [M. Guzman, G. Velasco, J. Castro and V. A. Zammit (1994) FEBS Lett. 344, 239-241].
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PMID:Insulin-independent and extremely rapid switch in the partitioning of hepatic fatty acids from oxidation to esterification in starved-refed diabetic rats. Possible roles for changes in cell pH and volume. 784 96

The maximal activity of the overt from of carnitine palmitoyltransferase I (CPT I; EC 2.3.1.21) and its sensitivity to inhibition by malonyl CoA were measured in mitochondria prepared from the livers of rats which had been fed for 10 weeks on either a low fat diet (LF; 2.4% fat by weight) or on one of four high fat diets which contained 20% by weight of either hydrogenated coconut oil (HCO), olive oil (OO), safflower oil (SO) or menhaden (fish) oil (MO). CPT I activity (i.e. activity per g of liver tissue), was elevated in animals fed the OO, SO or MO diets compared with those fed the LF or HCO diets. Feeding the HCO diet did not result in elevation of CPT I activity compared with feeding the LF diet. CPT I specific activity (i.e. activity per mg mitochondrial protein) was elevated in animals fed SO diet, but not in animals fed any of the other high fat diets. These observations suggest that an elevated fat load is not solely responsible for increasing CPT I activity, but that the fatty acid composition of the diet also plays a role. Hepatic CPT I activity of rats fed the LF diet was most sensitive to inhibition by malonyl CoA ([I50] = 0.53 microM). Each of the high fat diets decreased the sensitivity of CPT I to inhibition by malonyl CoA; CPT I activity in the livers from animals fed the MO diet was the least sensitive to malonyl CoA inhibition ([I50] = 1.8 microM). The fatty acid compositions of the major mitochondrial membrane phospholipids, phosphatidylcholine, phosphatidylethanolamine and cardiolipin were modified according to the fatty acid composition of the diet. Each of these phospholipids had a distinct fatty acid composition and similar effects of dietary lipid manipulation on the fatty acid compositions were observed. Feeding the SO diet resulted in fatty acid compositions which were most similar to those found after feeding the LF diet. Feeding the HCO and OO diets increased the proportions of stearic and oleic acids, respectively, while decreasing the proportion of linoeic acid. Feeding the MO diet resulted in increased proportions of palmitic, palmitoleic, eicosapentaenoic and docosahexaenoic acids and decreased proportions of linoleic and arachidonic acids in each of the phospholipids. It is proposed that the effects of dietary lipid manipulation upon CTP I activity and sensitivity to inhibition by malonyl CoA are due to alterations in the fatty acid composition of the phospholipids in the mitochondrial membrane where CPT I resides.
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PMID:The effect of dietary lipid manipulation on hepatic mitochondrial phospholipid fatty acid composition and carnitine palmitoyltransferase I activity. 786 92

Using the cDNA for rat liver mitochondrial carnitine palmitoyltransferase I (CPT I; EC 2.3.1.21) as a probe, we isolated its counterpart as three overlapping clones from a human liver cDNA library. Both the nucleotide sequence of the human cDNA and the predicted primary structure of the protein (773 aa) proved to be very similar to those of the rat enzyme (82% and 88% identity, respectively). The CPT I mRNA size was also found to be the same (approximately 4.7 kb) in both species. Screening of a human genomic library with the newly obtained cDNA yielded a positive clone of approximately 6.5 kb which, upon partial analysis, was found to contain at least two complete exons linked by a 2.3-kb intron. Oligonucleotide primers specific to upstream and downstream regions of one of the exon/intron junctions were tested in PCRs with DNA from a panel of somatic cell hybrids, each containing a single human chromosome. The results allowed unambiguous assignment of the human liver CPT I gene to the q (long) arm of chromosome 11. Additional experiments established that liver and fibroblasts express the same isoform of mitochondrial CPT I, legitimizing the use of fibroblast assays in the differential diagnosis of the "muscle" and "hepatic" forms of CPT deficiency. The data provide insights into the structure of a human CPT I isoform and its corresponding gene and establish unequivocally that CPT I and CPT II are distinct gene products. Availability of the human CPT I cDNA should open the way to an understanding of the genetic basis of inherited CPT I deficiency syndromes, how the liver CPT I gene is regulated, and which tissues other than liver express this particular variant of the enzyme.
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PMID:Human liver mitochondrial carnitine palmitoyltransferase I: characterization of its cDNA and chromosomal localization and partial analysis of the gene. 789 12

In newborn-pig hepatocytes, the rate of oleate oxidation is extremely low, despite a very low malonyl-CoA concentration. By contrast, the sensitivity of carnitine palmitoyltransferase (CPT) I to malonyl-CoA inhibition is high, as suggested by the very low concentration of malonyl-CoA required for 50% inhibition of CPT I (IC50). The rates of oleate oxidation and ketogenesis are respectively 70 and 80% lower in mitochondria isolated from newborn-pig liver than from starved-adult-rat liver mitochondria. Using polarographic measurements, we showed that the oxidation of oleoyl-CoA and palmitoyl-L-carnitine is very low when the acetyl-CoA produced is channelled into the hydroxymethylglutaryl-CoA (HMG-CoA) pathway by addition of malonate. In contrast, the oxidation of the same substrates is high when the acetyl-CoA produced is directed towards the citric acid cycle by addition of malate. We demonstrate that the limitation of ketogenesis in newborn-pig liver is due to a very low amount and activity of mitochondrial HMG-CoA synthase as compared with rat liver mitochondria, and suggest that this could promote the accumulation of acetyl-CoA and/or beta-oxidation products that in turn would decrease the overall rate of fatty acid oxidation in newborn- and adult-pig livers.
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PMID:Hepatic ketogenesis in newborn pigs is limited by low mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase activity. 790 71

Levels of mRNA for mitochondrial 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) synthase, carnitine palmitoyltransferase I (CPT I) and carnitine palmitoyltransferase II (CPT II), fatty acid synthase (FAS) and actin were analysed during liver regeneration. mRNA levels for mitochondrial HMG-CoA synthase decreased rapidly, reaching a minimum 12 h after partial hepatectomy and returning to normal at 24-36 h. In contrast, CPT I, CPT II and FAS mRNAs increased throughout the period examined. Expression of actin increased significantly during regeneration. Levels of mRNA for mitochondrial HMG-CoA synthase also decreased as a result of surgical stress, although the effect of hepatectomy was much greater. We determined the levels of mitochondrial HMG-CoA synthase using specific antibodies. The amount of protein rapidly decreased, although less markedly than the corresponding mRNA levels. These results show that the decrease described in ketogenesis in partially hepatectomized rats correlated with the decrease in the expression of mitochondrial HMG-CoA synthase, suggesting that this enzyme may also be a control point in ketogenesis in the regenerating liver, as it is in normal and diabetic rats.
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PMID:Gene expression of enzymes regulating ketogenesis and fatty acid metabolism in regenerating rat liver. 790 32


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