Mitochondrial fatty acid elongation

Mitochondrial fatty acid elongation occurs primarily when the [NADH]/[NAD+] ratio is high (e.g., anaerobiosis, excessive ethanol oxidation). 

When subcellular fractions of rabbit aorta were isolated, the mitochondria were found to be the major site of fatty acid synthesis. Mitochondria from atherosclerotic aortas incorporated acetate into fatty acids four times faster than did mitochondria from control aortas (Whereat, 1966). Cholesterol feeding in some way led to an acceleration of the mitochondrial fatty acid elongation pathway. 

A second system for fatty acid elongation exists in the mitosol, probably for provision of long fatty acids for mitochondrial structure. This system uses most of the same activities of 9-oxidation, but an NADPH dependent Enoyl-CoA reductase replaces the FAD dependent dehydrogenase. 

Numerous experiments were then performed with mitochondria incubated with acetate, CoA, ATP, etc., in attempts to detect fatty acid synthesis. In 1957, Lynen and his colleagues reported the presence in mitochondria of a system which catalyzed the elongation of caproyl CoA to octanoyl CoA by the addition of an acetate unit. NADH and NADPH had to be present. The existence of this mitochondrial system was confirmed by Wakil et al. in 1961 who showed the 12C acid could be extended to 16C by successive additions of 2C fragments. 

Pantothenic acid has a central role in energy-yielding metabolism as the functional moiety of coenzyme A (CoA), in the biosynthesis of fatty acids as the prosthetic group of acyl carrier protein, and through its role in CoA in the mitochondrial elongation of fatty acids the biosynthesis of steroids, porphyrins, and acetylcholine and other acyl transfer reactions, including postsynthetic acylation of proteins. Perhaps 4% of all known enzymes utilize CoA derivatives. CoA is also bound by disulfide links to protein cysteine residues in sporulating bacteria, where it may be involved with heat resistance of the spores, and in mitochondrial proteins, where it seems to be involved in the assembly of active cytochrome c oxidase and ATP synthetase complexes. 

The enzyme is present in two different forms. A membrane-bound 300 amino acid form is located mainly in the endoplasmic reticulum and outer mitochondrial membrane. The membrane form has a role in desaturation and elongation of fatty acids, cholesterol biosynthesis, and drug metabohsm. 

Mitochondrial elongation occurs by successive addition and reduction of acetyl units in a reversal of fatty acid oxidation. Although fatty acid P-oxidation and mitochondrial chain elongation have the same organelle location, reversal of a tra/ii-2-enoyl-CoA reductase P-oxidation is not feasible the FAD-dependent acyl-CoA dehydrogenase of P-oxidation is substituted by a more thermodynamically favorable enzyme reaction, catalyzed by NADPH-dependent enoyl-CoA reductase, to produce overall negative free-energy for the sequence. Enoyl-CoA reductase firom liver mitochondria is distinct from... 

Oxoglutarate can also serve as a starter piece for elongation by the oxoacid pathway. Extension by three carbon atoms yields 2-oxosuberate (Eq. 21-1). This dicarboxylate is converted by reactions shown in Eq. 24-39 into biotin and in archaebacteria into the coenzyme 7-mercaptoheptanoylthreonine phosphate (HTP), Eq. 21-1. Lipoic acid is also synthesized from a fatty acid, the eight-carbon octanoate. A fatty acid synthase system that utilizes a mitochondrial ACP may have as its primary fimction the synthesis of ocfanoate for lipoic acid formation. The mechanism of insertion of the two sulfur atoms to form lipoate (Chapter 15) is imcerfain. If requires an iron-sulfur protein jg probably similar to the corresponding process in the synthesis of biotin (Eq. 24-39)9 93a formation of HTP (Eq. 21-1). One component of the archaebacterial cofactor methano-furan (Chapter 15) is a tetracarboxylic acid that is formed from 2-oxoglufarafe by successive condensations with two malonic acid imits as in fatty acid synthesis. ... 

An unusual sphingolipid contains a 22-carbon, polyunsaturated fatty acid called du-panodonic acid, or 7,10,13,16,19-docosapentaenoic acid. In mammals, both the mitochondrial and endoplasmic reticular acyl-chain elongation and desaturation systems can synthesize clupanodonate from linolenate. 

Fig.1. Possible pathways for the intracellular movement of n-3 polyunsaturated fatty acid as it relates to the synthesis of 4,7,10,13,16,19-22 6. The pathway implies that when 24 6 (n-3) is produced in the endoplasmic reticulum, it preferentially moves to another cellular compartment rather than serving as a substrate for further chain elongation. It is not known whether fatty acids move between subcellular compartments as acyl-CoA or whether they are hydrolyzed followed by their reactivation at the subcellular site where they are to be metabolized. If 24 6 n-3 is to be metabolized by mitochondria, it must be transported across the outer (O.M.) and inner (I.M.) membranes into the mitochondrial matrix. This pathway has recently been shown to be of minor importance. The preferred, if not the exclusive pathway for 24 6 n-3 metabolism requires its movement to peroxisomes, where after one degradative cycle, the 22 6 n-3 preferentially moves back to the endoplasmic reticulum rather than serving as a substrate for continued (3-oxidation. Again it is not known in what form the 22 6 n-3 is transported, i.e., acyl-CoA or free fatty acid and how or whether these intracellular fatty acid movements require specific proteins.

A mitochondrial system for elongation of fatty acid chains, using acetyl-CoA as the two-carbon donor does exist but has limited activity with acyl-CoA substrates with 16 or more carbon atoms and is probably concerned with the lengthening of shorter chains. 

Examination of the fatty acid composition of brain microsomal and mitochondrial lipids (Table 2) reveals that major fatty acids of both particles are 16 0, 18 1, 20 4 and 22 6. It is not too surprising, then that under the influence of enzyme specificity, endogenous 20 4 should be most readily elongated (to 22 4). How-... 

In an attempt to locate such a pool of substrate fatty acid, we followed the incorporation of radioactivity from malonyl-or acetyl-CoA into both lipids and fatty acids of microsomal and mitochondrial preparations. As can be seen in Fig. 2, there is a hint that with the mitochondria the elongation products of the added substrate (18 2) follow the neutral lipids while 22 4, the... 

If this is true, it may be a difficult matter experimentally to test the brain elongation systems with added substrates. Nevertheless, kinetic studies have been made using the mitochondrial and microsomal elongation systems with several fatty acids. The Kju for both linoleic and linolenic acids with both microsomes and mitochondria was about 3 x 10 M while the V ax both substrates was somewhat greater for mitochondria than for microsomes and greater for 18 3 than for 18 2 (Fig. 4), For palmitate, under the same conditions, was 2 x 10 M and Vmax again somewhat... 

Besides the two classical mechanisms of saturated fatty acid synthesis, i.e., by condensation (cytoplasmic synthesis) and by elongation (mitochondrial synthesis), a mechanism of synthesis in micro-somes has recently been postulated. 

Howard (1968b) studied fatty acid synthetic systems in cell-free preparations from squirrel monkey aortas, and the data were similar to those for the rabbit aorta with regard to the mitochondrial system. Acetate or acetyl-CoA was a more efficient precursor than malonyl-CoA, and the Schmidt degradation data indicated that it was primarily an elongation system. The cytosol or HSS was examined, and malonyl-CoA was found to be incorporated into fatty acids 55-200 times more actively than acetyl-CoA, a finding that had been noted previously in liver HSS by Abraham et al. (1962a). Majerus and Lastra (1967) noted that malonyl-CoA was incorporated into fatty acids six or seven times as fast as acetyl-CoA by human leukocytes. The latter authors were unable to find any acetyl-CoA carboxylase activity in leukocytes and reasoned that these cells possess only the fatty acid synthetase. As they pointed out, in the absence of any acetyl-CoA carboxylase, the synthetase alone uses 1 mole of acetyl-CoA plus 7 moles of malonyl-CoA to make 1 mole of palmitate (Wakil... 

Fig. 1. The above pathways for fatty acid synthesis have been demonstrated to be present in the aorta. The thickness of the arrows denotes the author s interpretation of the relative contribution to total synthesis made by the three intracellular sites. The mitochondrial pathway has the largest capacity to utilize acetate for the elongation of available acyl units. The latter are derived from plasma free fatty acid (FFA) and lipolysis of tissue triglyceride (TG). The cytosol has a limited capacity to synthesize fatty acids from acetate because of minimal acetyl-CoA carboxylase (ACC) activity. The significance of fatty acid synthetase (FAS) activity is dubious in the absence of a source of malonyl-CoA. A microsomal elongation-desaturation pathway can synthesize a spectrum of saturated (SAT) and unsaturated (UNSAT) long-chain fatty acids, similar to the products of the mitochondrial system.

The product of the cytosol system probably is palmitic acid regardless of the tissue of origin. The microsomal and mitochondrial systems elongate available acyl units and, therefore, the products are multiple and variable. The synthesized fatty acids have many pathways available to them, and these have been discussed in volume 8 (cf. Portman). 

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