A4-Desaturase

FIGURE 3-7 Pathways for the interconversion of brain fatty acids. Palmitic acid (16 0) is the main end product of brain fatty acid synthesis. It may then be elongated, desaturated, and/or P-oxidized to form different long chain fatty acids. The monoenes (18 1 A7, 18 1 A9, 24 1 A15) are the main unsaturated fatty acids formed de novo by A9 desaturation and chain elongation. As shown, the very long chain fatty acids are a-oxidized to form a-hydroxy and odd numbered fatty acids. The polyunsaturated fatty acids are formed mainly from exogenous dietary fatty acids, such as linoleic (18 2, n-6) and a-linoleic (18 2, n-3) acids by chain elongation and desaturation at A5 and A6, as shown. A A4 desaturase has also been proposed, but its existence has been questioned. Instead, it has been shown that unsaturation at the A4 position is effected by retroconversion i.e. A6 unsaturation in the endoplasmic reticulum, followed by one cycle of P-oxidation (-C2) in peroxisomes [11], This is illustrated in the biosynthesis of DHA (22 6, n-3) above. In severe essential fatty acid deficiency, the abnormal polyenes, such as 20 3, n-9 are also synthesized de novo to substitute for the normal polyunsaturated acids. 

Guiet et al. (2003) demonstrated that deuterium (2H) distribution in fatty acids was non-statistical and could be related to isotopic discrimination during chain extension and desaturation. Petroselinic acid (C18 1A6) (Fig. 21.4), a fatty acid characteristic of the seeds of the Apiaceae, has been shown to be biosynthesized from palmitoyl-ACP (C16 0) by two steps, catalysed by a dedicated A4-desaturase and an elongase. The isotopic profile resulting from this pathway is similar to the classical plant fatty acid pathway, but the isotopic fingerprint from both the desaturase and elongase steps shows important differences relative to oleic and linoleic acid biosynthesis. 

Voss A, Reinhart M, Sankarappa S, Sprecher H. The metabolism of 7,10,13,16,19-docosapentaenoic acid to 4,7,10,13,16,19-docosahexaenoic acid inrat liveris independent of a4-desaturase. J Biol Chem 1991 266 19,995-20,000. 

The desaturation of fatty acids is usually assayed by incubating radioactive fatty acids with microsomes in the presence of appropriate cofactors. This general protocol has been used by many investigators to assay A9, A6, and A5 desaturase activities. It was thus assumed that 7,10,13,16-22 4 and 7,10,13,16,19-22 5 were desaturated by a microsomal A4 desaturase. In 1991, we showed that when rat liver microsomes were incubated with [1- C]7,10,13,16,19-22 5, it was not desaturated to 4,7,10,13,16,19-22 6. However, when malonyl-CoA was included in the incubation, the substrate was chain elongated to 9,12,15,18,21-24 5, which was then desaturated, at position-6, to yield 6,9,12,15,18,21-24 6. When [1- C]7,10,13,16,19-22 5 and the two [3- C]-labeled 24-carbon acids were incubated with hepatocytes, all three acids were metabolized to esterified [1- C]4,7,10, 13,16,19-22 6 (19). The findings implied that 4,7,10,13,16, 19-22 6 was made from 9,12,15-18 3 as follows ... 

The above-mentioned smdies raised several questions, the first of which is the following Do some cells express a A4 desaturase and thus not require a peroxisomal step to synthesize 4,7,10,13,16,19-22 6 Moore et al. (23) reported that [1- C] 6,9,12-18 3, [3-> C]7,10,13,16,19-22 5, [3- t]9,12,15,18, 21-24 5, and [3- C]6, 12,15,18,21-24 6 were all metabolized to 4,7,10,13,16,19-22 6 by control human skin fibroblasts, but not by fibroblasts from patients with Zellweger syndrome. Indeed, with cells from patients with Zellweger syndrome, labeled 6,9,12,15,18,21-24 6 accumulated as an end product, thus establishing the requirement of peroxisomes for the synthesis of 4,7,10,13,16,19-22 6. The labeling patterns of metabolites obtained when labeled n-3 acids were incubated with astrocytes (24) and retina (25) are also consistent with a pathway devoid of a A4 desaturation step. 

Elongation of arachidonic acid to 22 4 (7, 10, 13, 16) and the A4 desaturation to 22 5 (4, 7, 10, 13, 16) occur in liver in a very limited extent but may be important in adrenals and testis (Ayala et al, 1973). Similar conversion to 20 5 (5, 8, 11, 14, 17) acid that belongs to a-linolenic family to acids of 22 carbons is important in brain and it also requires a A4 desaturation. The A4 desaturase is very little active (Bernert and Sprecher, 1975 Ayala et al, 1973) and might be in consequence a second pass-marker. 

HTC cells (designated HTC for hepatoma tissue culture) were derived from the ascites form of a rat-carried Morris hepatoma 7288 C (Thompson et al, 1966). Previous studies have revealed that cells of this kind are able to desaturate and elongate fatty acids In this respect it was demonstrated that culture HTC cells preserved the ability to desaturate stearic to oleic acid ( A9 desaturase), a-linolenic acid to octdeca-6,9,12,15-tetraenoic acid (A6 desaturase), and eicosa-8,ll,14-trienoic acid to arachidonic acid (A5 desaturase) (Alaniz et al, 1975). They are also able to convert a-linolenic acid to higher homologs with 5 and 6 double bonds by desaturation and elongation reactions. These results also proved the existence of A4 desaturase activity (Alaniz et al, 1975). 

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