NADPH fatty acid synthase

One of the sex pheromone components of the housefly, Musca domestica, is Z9-21 H that is found on the cuticular surface of the fly. This compound is formed by the elongation of Z9-18 CoA using malonyl-CoA and NADPH to Z15-24 CoA which is decarboxylated to form Z9-21 Hc (Fig. 3) [78-80]. Other pheromone components include an epoxide and ketone that are produced from Z9-21 Hc by a cytochrome P450 [81,82] and methyl-branched alkanes that are produced by the substitution of methylmalonyl-CoA in place of malonyl-CoA at specific points during chain elongation [83,84]. A novel microsomal fatty acid synthase is involved in production of methyl-branched alkanes in most insects [85-87]. This fatty acid synthase is different from the ubiquitous soluble fatty acid synthase that produces saturated straight chain fatty acids in that it is found in the microsomes and prefers methylmalonyl-CoA. The amino acids valine and isoleucine can provide the carbon skeletons for methylmalonyl-CoA as well as propionate [83]. 

Rate-limiting Citrate activates, insulin activates Fatty acid synthase (requires NADPH)... 

Figure 11.4 Condensation, dehydration and reduction reactions in fatty add synthesis. These reactions constitute the major components of the pathway of fatty acid synthesis and are all catalysed by fatty acid synthase. The reduction reactions, indicated by addition of 2H in the diagram, involve the conversion of NADPH to NADP . (The re-conversion of NADP back to NADPH occurs in the pentose phosphate pathway.) The condensation reaction results in an increase in size of acyl-ACP by two carbon units in each step. The two carbons for each extension are each provided by malonyl-CoA. ACP - acyl carrier protein.

The first step is carboxylation of acetyl CoA to malonyl CoA. This reaction is catalyzed by acetyl-CoA carboxylase [5], which is the key enzyme in fatty acid biosynthesis. Synthesis into fatty acids is carried out by fatty acid synthase [6]. This multifunctional enzyme (see p. 168) starts with one molecule of ace-tyl-CoA and elongates it by adding malonyl groups in seven reaction cycles until palmi-tate is reached. One CO2 molecule is released in each reaction cycle. The fatty acid therefore grows by two carbon units each time. NADPH+H is used as the reducing agent and is derived either from the pentose phosphate pathway (see p. 152) or from isocitrate dehydrogenase and malic enzyme reactions. 

The rest of the steps in fatty acid synthesis are catalyzed by the fatty acid synthase com plex, which produces palmitoyl CoA from acetyl CoA and malonyl CoA, with NADPH as the source of reducing equivalents. 

Overview Fatty acid synthesis involves the condensation of two-carbon units, in the form of acetyl CoA, to form long hydrocarbon chains in a series of reactions. These reactions are carried out on the fatty acid synthase complex using NADPH as reductant. The fatty acids are covalently linked to acyl carrier protein (ACP) during their synthesis. 

Figure 2.5. Reaction sequence for the biosynthesis of fatty acids de novo by the animal FAS. The condensation reaction proceeds with stereochemical inversion of the malonyl C-2, the (3-ketoacyl moiety is reduced by NADPH to D-(3 hydroxyacyl moiety, which then is dehydrated to a trans-enoyl moiety finally, the enoyl moiety is reduced to a saturated acyl moiety by NADPH, with the simultaneous addition of a solvent proton. The two C atoms at the methyl end of the fatty acid are derived from acetyl-CoA, the remainder from malonyl-CoA. The entire series of reactions takes approximately 1 second. PSH, phosphopantetheine. (Reprinted from Prog, in Lipid Res., vol. 42, S. Smith, A. Witkowski and A.K. Joshi, Stuctural and functional organization of the animal fatty acid synthase, pp. 289-317, copyright (2003), with permission from Elsevier).

The enzyme system that catalyzes the synthesis of saturated long-chain fatty acids from acetyl CoA, malonyl CoA, and NADPH is called the fatty acid synthase. The constituent enzymes of bacterial fatty acid synthases are dissociated when the cells are broken apart. The availability of these isolated enzymes has facilitated the elucidation of the steps in fatty acid synthesis (Table 22.2). In fact, the reactions leading to fatty acid synthesis in higher organisms are very much like those of bacteria. 

This four-step cycle includes condensation of acetate and malonate to give ketobu-tanoate with subsequent reduction to butanoate in three further steps. These are reduction to the 3R hydroxy acid, dehydration to the 2t acid, and reduction again. Reduction is affected by NADPH and a proton. The process is then repeated to add further two-carbon units until a thioesterase liberates the free acid. This sequence requires a fatty acid synthase, which contains the enzymes needed for each of the four steps viz. p-ketoacyl-ACP synthase, p-ketoacyl-ACP reductase, p-ketoacyl-ACP dehydrase, and enoyl-ACP reductase, respectively. 

Elongation microsomal process (ER), requires malonylCoA and two NADPH, process is similar to that occurring with fatty acid synthase. 

Regulation of NADPH Production Fatty Acid Synthase Respiratory Quotient Data from Human Studies... 

The growing fatty acid chain on the fatty acid synthase complex is elongated, two carbons at a time, by the addition of the three-carbon compound, malonyl CoA, which is subsequently decarboxylated. With each two-carbon addition, the growing chain, which initially contains a P-keto group, is reduced in a series of steps that require NADPH. NADPH is produced by the pentose phosphate pathway and by the reaction catalyzed by the malic enzyme. 

NADPH supplies reducing equivalents for reactions that occur on the fatty acid synthase complex. 

B. The synthesis of fatty acids from glucose occurs in the cytosol, except for the mitochondrial reactions in which pyruvate is converted to citrate. Biotin is required for the conversion of pyruvate to oxaloacetate, which combines with acetyl CoA to form citrate. Biotin is also required by acetyl CoA carboxylase. Pantothenic acid is covalently bound to the fatty acid synthase complex as part of a phosphopantetheinyl residue. The growing fatty acid chain is attached to this residue during the sequence of reactions that produce palmitic acid. NADPH, produced by the malic enzyme as well as by the pentose phosphate pathway, provides reducing equivalents. Citrate, not isocitrate, is a key regulatory compound. 

Cytoplasmic fatty acid synthase yields palmitate. Human triacylglycerol contains fatty acids with 18, 20, 22, and 24 carbon atoms, which are synthesized by elongation of palmitate in endoplasmic reticulum or mitochondria. Elongation in the endoplasmic reticulum occurs mainly in liver and involves Cio-ie-saturated and Cig-unsaturated fatty acids by successive addition of two-carbon groups derived from malonyl-CoA (Figures 18-14 and 18-15). The reductant is NADPH. The intermediates, however, are CoA thioesters. 

The first step in de novo fatty acid synthesis is the production of malonyl-CoA from acetyl-CoA and bicarbonate. This committed step is catalyzed by acetyl-CoA carboxylase present in the cytoplasm of liver cells and adipocytes. After replacement of the CoA residue in acetyl-CoA by ACP (acyl carrier protein), malonyl-ACP is used to convert acetyl-ACP to butyryl-ACP by the fatty acid synthase complex. In this multistep reaction, NADPH is used as donor of hydrogen atoms and CO2 is produced. Butyryl-ACP is subsequently elongated to hexanoyl-ACP by a similar process in which malonyl-ACP serves as donor of two carbon atoms required for lengthening of the growing acyl chain. This process is repeated until palmitic acid... 

Triketides are relatively rare. Triacetic acid lactone (4.2) has been detected in Penicillium patulum. It is also produced by fatty acid synthase in the absence of the reductant NADPH. Radicinin (4.3) is a major phytotoxin isolated from Ahernaria radicina (Stemphyllium radicinum) which causes a black rot of carrots. It is also formed by other Ahernaria species. Its pyrano[4,3- ]pyran structure, the identification of which had eluded purely chemical degradative studies, was established in one of the earlier applications of NMR spectroscopy to natural product structure elucidation. The biosynthesis of radicinin from acetate units was studied in 1970 by both radio-isotope methods using carbon-14 and by carbon-13 enrichment studies with NMR methods of detection. This was one of the first applications of this NMR technique to biosynthetic problems. These results established the labelling pattern for radicinin shown in 4.3. 

Fatty acid biosynthesis occurs by the sequential addition of acetyl groups and, on first inspection, appears to be a simple reversal of the (3-oxidation pathway. Although the biochemical reactions are similar, fatty acid synthesis differs from (3-oxidation in the following ways It occurs in the cytoplasm, utilizes acyl carrier protein and NADPH, and is carried out by a multienzyme complex, fatty acid synthase. 

Malonyl-CoA is then used for fatty acid synthesis. This process requires Fatty Acid Synthase (FAS) (Chirala and Wakil, 2004) that uses acetyl-CoA as a primer, malonyl-CoA as a two-carbon donor, and NADPH as a reducing equivalent (Fig. 1.1). The predominant fatty acid produced by FAS is palmitic acid (Cl 6 0). The structure of FAS has been extensively studied (Asturias et al., 2005 Maier et ah, 2006). FAS is a multifunctional complex consisting of two identical monomers. However, only the dimeric form is active (Chirala et al.,... 

Abbreviations FASN, fatty acid synthase ACC, acetyl-CoA-carboxylase ACL, ATP-citrate lyase NADPH, nicotinamide adenine dinucleotide phosphate MAT, malonyl acetyl transferases KS, ketoacyl synthase KR, p-ketoacyl reductase DH, p-hydroxyacyl dehydratase ER, enoyl reductase TE, thioesterase ACP, acyl carrier protein VLCFA, very long chain fatty acids ELOVL, elongation of very long chain fatty acids SCDl, stearoyl-CoA desaturase-1 AMPK, AMP-activated kinase ME, malic enzyme FASKOL, liver-specific deletion of FAS PPARa, Peroxisome Proliferator-Activating Receptor alpha HMG-CoA, 3-hydroxy-3-methyl-glutaryl-CoA SREBP, sterol response element binding protein SIP, site-one protease S2P, site-two... 

Fig. 8. Pathways involved in the conversion of glucose to fatty acid. Reaction (1) is catalyzed by cytosolic malate dehydrogenase. Reaction (2) is catalyzed by mitochondrial malate dehydrogenase. (T) designates tricarboxylate anion transporter. Reactions catalyzed by glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase in the pentose phosphate pathway produce NADPH. CS, citrate synthase ACL, ATP citrate lyase PDH, pyruvate dehydrogenase complex ACC, acetyl-CoA carboxylase FAS, fatty acid synthase.

Adipocytes readily convert the products of glycolysis into fatty acids via the de novo biosynthetic pathway (Chapter 6). Briefly, surplus citrate is transported from the mitochondrion and cleaved to produce cytosolic acetyl-CoA. This acetyl-CoA is acted upon by acetyl-CoA carboxylase producing malonyl-CoA. The next steps of the fatty acid biosynthetic pathway are carried out by the multifunctional fatty acid synthase that utilizes NADPH to catalyze multiple condensations of malonyl-CoA with acetyl-CoA or the elongating lipid, eventually generating palmitate. 

The growing fatty acid chain, attached to the fatty acid synthase complex in the cytosol, is elongated by the sequential addition of 2-carbon units provided by malonyl CoA. NADPH, produced by the pentose phosphate pathway and the malic enzyme, provides reducing equivalents When the growing fatty acid chain is 16 carbons in length, it is released as palmitate After activation to a CoA derivative, palmitate can be elongated and desaturated to produce a series of fatty acids. 

As an overview, fatty acid synthase sequentially adds 2-carbon units from malonyl CoA to the growing fatty acyl chain to form palmitate. After the addition of each 2-carbon unit, the growing chain undergoes two reduction reactions that require NADPH. 

Fig. 36.4. Regulation of citrate lyase, malic enzyme, glucose 6-phosphate dehydrogenase, and fatty acid synthase. Citrate lyase, which provides acetyl CoA for fatty acid biosynthesis, the enzymes that provide NADPH (malic enzyme, glucose 6-phosphate dehydrogenase), as well as fatty acid synthase, are inducible (circled t).

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