Fatty acyl-CoA derivatives

All of the other enzymes of the /3-oxidation pathway are located in the mitochondrial matrix. Short-chain fatty acids, as already mentioned, are transported into the matrix as free acids and form the acyl-CoA derivatives there. However, long-chain fatty acyl-CoA derivatives cannot be transported into the matrix directly. These long-chain derivatives must first be converted to acylearnitine derivatives, as shown in Figure 24.9. Carnitine acyltransferase I, located on the outer side of the inner mitochondrial membrane, catalyzes the formation of... 

Lynen had studied chemistry in Munich under Wieland his skill as a chemist led to the successful synthesis of a number of fatty acyl CoA derivatives which proved to be substrates in the catabolic pathway. Many of these C=0 or C=C compounds had characteristic UV absorption spectra so that enzyme reactions utilizing them could be followed spectrophotometrically. This technique was also used to identify and monitor the flavoprotein and pyridine nucleotide-dependent steps. Independent evidence for the pathway was provided by Barker, Stadtman and their colleagues using Clostridium kluyveri. Once the outline of the degradation had been proposed the individual steps of the reactions were analyzed very rapidly by Lynen, Green, and Ochoa s groups using in the main acetone-dried powders from mitochondria, which, when extracted with dilute salt solutions, contained all the enzymes of the fatty acid oxidation system. 

The condensation of acetyl CoA and oxaloacetate to form citrate is catalyzed by citrate synthase (Figure 9.5). This aldol condensation has an equilibrium far in the direction of citrate synthesis. Citrate synthase is allosterically activated by Ca2+ and ADP, and inhibited by ATP, NADH, succinyl CoA, and fatty acyl CoA derivatives (see Figure 9.9). However, the primary mode of regulation is also deter mined by the availability of its substrates, acetyl CoA and oxaloac etate. [Note Citrate, in addition to being an intermediate in the TCA cycle, provides a source of acetyl CoA for the cytosolic synthesis of... 

Regulation of the LDL receptor gene involves a hormone-response element (HRE, see p. 238).] Third, if the cholesterol is not required immediately for some structural or synthetic purpose, it is esterified by acyl CoA cholesterol acyltransferase (ACAT, AC AT transfers a fatty acid from a fatty acyl CoA derivative to cholesterol, producing a cholesteryl ester that can be stored in the cell (Figure 18.21). The activity of ACAT is enhanced in the presence of increased intracellular cholesterol. 

Citrate is synthesized from oxaloacetate (OAA) and acetyl CoA by citrate synthase. This enzyme is allosterically activated by ADP, and inhibited by ATP, NADH, succinyl CoA, and fatty acyl CoA derivatives. Citrate is isomerized to isocitrate by aconitase, an enzyme that is targeted by the rat poison, fluoroacetate. 

Fatty acyl-CoA derivatives 507 Fatty alcohols 380-382,382 Favin 64s... 

Acetyl CoA Carboxylase Acetyl CoA carboxylase catalyzes the first and rate-Umiting step of fatty acid synthesis carboxylation of acetyl CoA to malonyl CoA. The mammalian enzyme is activated allostericaUy by citrate and isocitrate, and inhibited by long-chain fatty acyl CoA derivatives. It is also activated in response to insulin and inactivated in response to glucagon. 

Short-chain fatty acyl CoA derivatives are inhibitors of pantothenate kinase in perfused rat hearts, the addition of any of the major energy-yielding... 

Short to medium length fatty acids are permeable to the mitochondrial membrane. They are activated to fatty acyl CoA derivatives in the mitochondrial matrix by Butyryl-CoA Synthetase ... 

The CoA-fortlfled microsomal enzyme system was inhibited by SKF 525-A (B-diethylaminoethyl-diphenylpropyl acetate) and by 2,4-dlchloro-6-phenylphenoxyethylamine HBr (DPEA) in a dose-dependent fashion (28). Both are known inhibitors of the classical hepatic microsomal mixed-function oxidase system. This inhibition has been suggested to Involve a nonspecific binding to liver microsomal proteins and/or phospholipids (18.19). Binding to phospholipids would support the hypothesis proposed by Swell and co-workers (10) that fatty acyl CoA derivatives were not necessarily the only immediate source for the fatty acid moieties of the conjugates that phospholipids may act as the fatty acids reservoir in the CoA-fortified enzyme system. However, it is also known that cholesterol esterification is very sensitive to the fluidity of the microsomal membrane (12). The reduced conjugation produced when... 

Fatty acyl CoA formation, like the phosphorylation of glucose, is a prerequisite to metabolism of the fatty acid in the cell (Fig. 23.3). The multiple locations of the long-chain acyl CoA synthetase reflects the location of different metabohc routes taken by fatty acyl CoA derivatives in the cell (e.g., triacylglycerol and phosphohpid synthesis... 

Carnitine serves as the carrier that transports activated long chain fatty acyl groups across the inner mitochondrial membrane (Fig. 23.4). Carnitine acyl transferases are able to reversibly transfer an activated fatty acyl group from CoA to the hydroxyl group of carnitine to form an acylcamitine ester. The reaction is reversible, so that the fatty acyl CoA derivative can be regenerated from the carnitine ester. 

Fatty acid oxidation also may be restricted by the mitochondrial CoASH pool size. Acetyl CoASH units must enter the TCA cycle or another metabolic pathway to regenerate CoASH required for formation of the fatty acyl CoA derivative from fatty acyl carnitine. 

This CoA derivative then exchanges its CoA for another partner called carnitine. The carnitine-fatty acid complex, which is soluble in the mitochondrial membranes, is then transported from the cytosol past the inner mitochondrial membrane into the inner matrix, where it exchanges the carnitine for acetyl-CoA, to again become the fatty acyl-CoA derivative. 

Problem 22.52. True or false The first step in the oxidation of a fatty acyl-CoA derivative requires... 

If each enzyme could operate only on fatty acyl CoA derivatives of a particular chain length, then as many as eight sets of enzymes would be required to carry out the (3 oxidation of palmitate. The lact that most enzymes of the (3-oxidation pathway can use acyl CoA molecules of different chain lengths as substrates means that the cell needs to synthesize fewer different enzymes to carry out fatty acid oxidation. 

It is the view of the present authors that a physiological role for these inhibitors cannot be disregarded on the basis of these arguments rather, the question must remain open until more definitive experiments are performed. It seems reasonable that, in the intracellular environment, the critical micelle concentration for the fatty acyl-CoA derivatives may never be achieved and that the largest fraction of these inhibitory substances is bound at hydrophobic sites of intracellular proteins and lipid structures such as membranes. In this case, the degree of inhibition would depend on the relative affinity of hydrophobic sites on the carboxylase, the concentration of free fatty acyl-CoA, and that bound at other intracellular hydrophobic sites. 

There is abundant evidence indicating that a natural hydrophobic inhibitor of acetyl-CoA carboxylase is present in crude enzyme extracts of liver and adipose tissue [128,129,182,192,236-238]. The activating effect of (+)-palmityl carnitine on fatty acid synthesis in crude liver extracts and on impure acetyl-CoA carboxylase preparations has tentatively been ascribed to the displacement of hydrophobic inhibitors such as fatty acids or fatty acyl-CoA derivatives [129,182,192,236-238]. Inhibition of rat liver acetyl-CoA carboxylase by added palmityl-CoA can be reversed in part by (+)-palmityl carnitine [236], but not by citrate. This activating effect does not appear to be specific with respect to (+)-palmityl carnitine in that cetyl trimethylammonium ion is also effective [192]. Furthermore, impure preparations of acetyl-CoA carboxylase from adipose tissue or rat liver are markedly activated by serum albumin [123,129,238] or extensive dilution of the enzyme preparation prior to assay [129,182]. On the other hand, none of these agents [(+)-palmityl carnitine, serum albumin, or dilution], which activate the impure carboxylase, have an activating effect on the homogeneous acetyl-CoA carboxylases from adipose tissue or liver [129,182, 239]. It is evident that an inhibitory substance, apparently hydrophobic in nature, is removed either by purification of the enzyme or by the agents or treatments mentioned above. 

It has been reported that, like liver acetyl-CoA carboxylase, both the liver and yeast fatty acid synthetases are inhibited by low concentrations (0.5 to 5 X 10 71/) of long-chain fatty acyl-CoA derivatives, the longer-chain derivative producing greater inhibition [226,246,247]. In the case of the yeast synthetase, inhibition by long-chain acyl-CoA derivatives was competitive with respect to acetyl-CoA and NADPH. For the same reasons alluded to earlier in the discussion of the inhibition of acetyl-CoA carboxylase by fatty acyl-CoA derivatives, some caution must be exercised in interpreting the effect of these potent inhibitors (see Section V, C, 2). 

Similarly the fatty acids must be activated by conversion to their CoA derivatives before they can be metabolized. Formation of the fatty acyl-CoA derivatives is catalysed by various/affy acid thiokinases (fatty acid CoA ligases) whose activity is linked with the breakdown of ATP to AMP and pyrophosphate, the liberated energy being used in the formation of the thiol ester bond ... 

There are three separate sets of enzymes catalysing these four reactions, with specificity for long-, medium- and short-chain fatty acyl CoA derivatives. Each set of enzymes is arranged as a membrane-bound array, and the product of one is passed directly to the active site of the next. The result of this is that, although short- and medium-chain fatty acyl CoA derivatives can be detected in the mitochondrial matrix, as they pass from one array of enzymes to the next, none of the intermediates of the reaction spiral can be detected - they remain enzyme bound. 

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