Carnitine formation

The most likely deficiency is a lack of 2,4-dienoyl CoA reductase, an enzyme that is essential for the degradation of unsaturated fatty acids with double bonds at even-numbered carbons. Such fatty acids include linoleate (9-ds,12-ds 18 2). Four rounds of oxidation of linoleoyl CoA generate a 10-carbon acyl CoA that contains a trans-A and a cis-A double bond. This intermediate is a substrate for the reductase, which converts the 2,4-dienoyl CoA to ds-A -enoyl CoA. A dehciency of 2,4-dienoyl reductase leads to an accumulation of trans-A, ds-A -decadienoyl CoA molecules in the mitochondrion. The observation that carnitine derivatives of the 2,4-dienoyl CoA are found in blood and urine provides evidence that these molecules accumulate in the mitochondrion and are then attached to carnitine. Formation of carnitine decadienoate allows the acyl molecules to be transported across the inner mitochondrial membrane into the cytosol, and then into the circulation. 

Detoxifica.tlon. Detoxification systems in the human body often involve reactions that utilize sulfur-containing compounds. For example, reactions in which sulfate esters of potentially toxic compounds are formed, rendering these less toxic or nontoxic, are common as are acetylation reactions involving acetyl—SCoA (45). Another important compound is. Vadenosylmethionine [29908-03-0] (SAM), the active form of methionine. SAM acts as a methylating agent, eg, in detoxification reactions such as the methylation of pyridine derivatives, and in the formation of choline (qv), creatine [60-27-5] carnitine [461-06-3] and epinephrine [329-65-7] (50). 

Biochemical Functions. Ascorbic acid has various biochemical functions, involving, for example, coUagen synthesis, immune function, dmg metabohsm, folate metaboHsm, cholesterol cataboHsm, iron metaboHsm, and carnitine biosynthesis. Clear-cut evidence for its biochemical role is available only with respect to coUagen biosynthesis (hydroxylation of prolin and lysine). In addition, ascorbic acid can act as a reducing agent and as an effective antioxidant. Ascorbic acid also interferes with nitrosamine formation by reacting direcdy with nitrites, and consequently may potentially reduce cancer risk. 

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... 

FIGURE 24.9 The formation of acylcar-nitines and their transport across the inner mitochondrial membrane. The process involves the coordinated actions of carnitine acyltrans-ferases on both sides of the membrane and of a translocase that shuttles O-acylcarnitines across the membrane. 

A number of iron-containing, ascorbate-requiring hydroxylases share a common reaction mechanism in which hydroxylation of the substrate is linked to decarboxylation of a-ketoglutarate (Figure 28-11). Many of these enzymes are involved in the modification of precursor proteins. Proline and lysine hydroxylases are required for the postsynthetic modification of procollagen to collagen, and prohne hydroxylase is also required in formation of osteocalcin and the Clq component of complement. Aspartate P-hydroxylase is required for the postsynthetic modification of the precursor of protein C, the vitamin K-dependent protease which hydrolyzes activated factor V in the blood clotting cascade. TrimethyUysine and y-butyrobetaine hydroxylases are required for the synthesis of carnitine. 

GMBS or sulfo-GMBS have been used for studying carnitine palmitoyltransferase-1 in its formation of a complex within the outer mitochondrial membrane (Faye et al., 2007), for investigating protein organization of the postsynaptic density (Liu et al., 2006), and in studying the structure and dynamics of rhodopsin (Jacobsen et al., 2006). 

Mitochondria contain all the enzymes necessary for oxidation of fatty acids but, before this can take place, the fatty acids have to be transported into the mitochondria. Transport requires the formation of an ester of the fatty acid with a compound, carnitine, in a reaction catalysed by the enzyme carnitine palmitoyltransferase ... 

Epididymis Carnitine Inositol Phosphatidylcholine Cholesterol Glycoproteins Facihtates acetyl-CoA oxidation by spermatozoa (Chapter 9) Precursor for formation of phosphatidyhnositol bisphosphate Buffer to maintain pH and a source of chohne Stabilises membranes They coat the surface of the sperm to protect against IgA... 

Pharmacology Vitamin C, a water-soluble vitamin, is an essential vitamin in man however, its exact biological functions are not fully understood. It is essential for the formation and the maintenance of intercellular ground substance and collagen, for catecholamine biosynthesis, for synthesis of carnitine and steroids, for conversion of folic acid to folinic acid and for tyrosine metabolism. 

Ascorbic acid or vitamin C is found in fruits, especially citrus fruits, and in fresh vegetables. Man is one of the few mammals unable to manufacture vitamin C in the liver. It is essential for the formation of collagen as it is a cofactor for the conversion of proline and lysine residues to hydroxyproline and hydroxylysine. It is also a cofactor for carnitine synthesis, for the conversion of folic acid to folinic acid and for the hydroxylation of dopamine to form norepinephrine. Being a lactone with two hydroxyl groups which can be oxidized to two keto groups forming dehydroascorbic acid, ascorbic acid is also an anti-oxidant. By reducing ferric iron to the ferrous state in the stomach, ascorbic acid promotes iron absorption. 

Mechanism of Action Assists in collagen formation and tissue repair and is involved in oxidation reduction reactions and other metabolicreactions.TAerapeMficEffect Involved in carbohydrate use and metabolism, as well as synthesis of carnitine, lipids, and proteins. Preserves blood vessel integrity. 

Under physiologic conditions, carnitine is primarily required to shuttle long-chain fatty acids across the inner mitochondrial membrane for FAO and products of peroxisomal /1-oxidation to the mitochondria for further metabolism in the citric acid cycle [40, 43]. Acylcarnitines are formed by conjugating acyl-CoA moieties to carnitine, which in the case of activated long-chain fatty acids is accomplished by CPT type I (CPT-I) [8, 44]. The acyl-group of the activated fatty acid (fatty acyl-CoA) is transferred by CPT-I from the sulfur atom of CoA to the hydroxyl group of carnitine (Fig. 3.2.1). Carnitine acylcarnitine translocase (CACT) then transfers the long-chain acylcarnitines across the inner mitochondrial membrane, where CPT-II reverses the action of CPT-I by the formation of acyl-CoA and release of free un-esterified carnitine. 

The incidence of the severe form is between 1 in 20,000 and 1 in 40,000 in adults, although the incidence is much higher in children (1 in 5000). The fatty liver is a "visible" symptom of dysfunction, not necessarily a cause of liver failure, although it can be. Valproic acid is similar to a fatty acid and therefore can become incorporated into fatty acid metabolism. This involves formation of an acyl CoA derivative and also a carnitine derivative. However, this depletes both CoA from the intramitochondrial pool and carnitine and so compromises the mitochondria and reduces the ability of the cell to metabolize short-, medium-, and long-chain fatty acids via p-oxidation (Fig. 7.15). 

Similarly, factors that stimulate acetyl-CoA carboxylase, the first enzyme in the pathway for fatty acid synthesis, also discourage fatty acid catabolism. This dual effect occurs because the first enzyme in the pathway leads to the formation of malonyl-CoA, which is a potent inhibitor of carnitine acyltransferase I. This inhibition prevents the transport of fatty acids into the mitochondrion, thereby, preventing fatty acid breakdown. 

The catabolism of lysine merges with that of tryptophan at the level of (3-ketoadipic acid. Both metabolic pathways are identical from this point on and lead to the formation of acetoacetyl-CoA (Figure 20.21). Lysine is thus ketogenic. It does not transaminate in the classic way. Lysine is a precursor of carnitine the initial reaction involves the methylation of e-amino groups of protein-bound lysine with SAM. The N-methylated lysine is then released proteolytically and the reaction sequence to carnitine completed. See Equation (19.6) for the structure of carnitine. 

Carnitine deficiency complicates HMG-CoA lyase deficiency and other inborn errors of metabolism, which results in organic acidemia. L-Camitine or P-hydroxy-y-trimethylammonium butyrate is a carrier molecule that transports long-chain fatty acids across the inner mitochondrial membrane for subsequent P-oxi-dation. L-Carnitine also facilitates removal of toxic metabolic intermediates or xenobiotics via urinary excretion of their acyl carnitine derivatives. Indeed, individuals with HMG-CoA lyase deficiency have been shown to excrete 3-methylgluatarylcamitine (Roe et al., 1986). In the absence of ketogenesis, the formation of the acyl carnitine derivative of 3-hydroxy-3-methylglutarate from HMG-CoA also serves to regenerate free CoA in the mitochondria and permits continued P-oxidation of fatty acids. 

Carnitine palmitoyltransferases I and II catalyze the transfer of long-chain acyl coenzyme A into mitochondria. The I isozyme is located on the cytosol side of the inner membrane and catalyzes the formation acylcarnitine from acyl-CoA and carnitine. After acylcarnitine crosses the inner membrane, it is converted back to acyl-CoA by the action of the II isozyme. This assay measures the activity of carnitine palmitoyltransferase I in intact mitochondria. 

As a result of the reduced activity of the mutase in vitamin B12 deficiency, there is an accumulation of methyhnalonyl CoA, some of which is hydrolyzed to yield methylmalonic acid, which is excreted in the urine. As discussed in Section 10.10.3, this can be exploited as a means of assessing vitamin B12 nutritional status. There may also be some general metabolic acidosis, which has been attributed to depletion of CoA because of the accumulation of methyl-malonyl CoA. However, vitamin B12 deficiency seems to result in increased synthesis of CoA to maintain normal pools of metabolically useable coenzyme. Unlike coenzyme A and acetyl CoA, neither methylmalonyl CoA nor propionyl CoA (which also accumulates in vitamin B12 deficiency) inhibits pantothenate kinase (Section 12.2.1). Thus, as CoA is sequestered in these metabolic intermediates, there is relief of feedback inhibition of its de novo synthesis. At the same time, CoA may be spared by the formation of short-chain fatty acyl carnitine derivatives (Section 14.1.1), which are excreted in increased amounts in vitamin B12 deficiency. In vitamin Bi2-deficient rats, the urinary excretion of acyl carnitine increases from 10 to 11 nmol per day to 120nmolper day (Brass etal., 1990). 

The major functions of pantothenic acid are in CoA (Section 12.2.1) and as the prosthetic group for AGP in fatty acid synthesis (Section 12.2.3). In addition to its role in fatty acid oxidation, CoA is the major carrier of acyl groups for a wide variety of acyl transfer reactions. It is noteworthy that a wide variety of metabolic diseases in which there is defective metabolism of an acyl CoA derivative (e.g., the biotin-dependent carboxylase deficiencies Sections 11.2.2.1 and 11.2.3.1), CoA is spared by formation and excretion of acyl carnitine derivatives, possibly to such an extent that the capacity to synthesize carnitine is exceeded, resulting in functional carnitine deficiency (Section 14.1.2). 

In general, the effects on collagen synthesis are more marked and more important than those of decreased formation of carnitine (as a result of impaired activity of trimethyllysine and y-butyrobetaine hydroxylases Section 14.1.1), impaired xenobiotic metabolism, or hypercholesterolemia (Section 13.3.8). However, depletion of muscle carititine may account for the lassitude and fatigue that precede clinical signs of scurvy. 

The total body content of carnitine is about 100 mmol, and about 5% of this turns over daily. Plasma total carnitine is between 36 to 83 /rmol per L in men and 28 to 75 /rmol per L in women, mainly as free carnitine. Although both free carnitine and acyl carnitine esters are excreted in the urine, much is oxidized to trimethylamine and trimethylamine oxide. It is not known whether the formation of trimethylamine and trimethylamine oxide is caused by endogenous enzymes or intestinal bacterial metabolism of carnitine. 

In general, the effects on collagen synthesis are more marked and more important than those of decreased formation of carnitine (as a result of impaired activity of trimethyllysine and y-butyrobetaine hydroxylases Section... 

Figure 30.9. Control of Fatty Acid Degradation. Malonyl Co A inhibits fatty acid degradation by inhibiting the formation of acyl carnitine.

Carnitine is required for transport of longoxidative metabolism as well as in the formation of ketone bcidies, The concentration of free carnitine in muscle is about 4,0 mmol/kg. The concentration of carnitine bound to long-chain fatty adds (fatty acyl-camitine) is lower, about 0,2 mmol/kg. Short-chain fatty adds, including acetic, are also esterified to carnitine, but the functions of these complexes are not clear. There is some indication that keto forms of BCAAs (BCKAs) can also be esterified to carnitine. These complexes can then be transported into the mitochondria for complete oxidation of the BCKAs, The importance of this mode of BCKA transport is not dear (Takakura et ai., 1997). 

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