BIOCHEMISTRY - Textbook - Ostapchenko L. I. - 2012

Chapter 14. LIPID METABOLISM

14.4. Biosynthesis of Higher Fatty Acids

Fatty acids are synthesized in the animal body predominantly during the absorptive state, mainly within the Cells of The Liver and adipose tissue. During this period, cells are generally fully supplied with the metabolic substrates, energy, and reducing equivalents required for de novo fatty acid synthesis in the form of acetyl-CoA, ATP, and NADPH, respectively.

The enzyme systems responsible for fatty acid synthesis are localized in both the Cytoplasm and the mitochondrial matrix; however, the cytoplasmic synthesis system significantly surpasses the mitochondrial one in activity, making the cytoplasm the primary site of FATTY ACID Biosynthesis.

The substrate for synthesis—acetyl-CoA molecules produced in the Cell/35.html">Mitochondria via the Oxidation of Pyruvate and fatty acids—is transported into the cytoplasm through several transport systems: in the form of acetate, citrate, and with the participation of carnitine.

When The Tricarboxylic Acid Cycle is inhibited by an excess of ATP, citrate (formed in the reaction between acetyl-CoA and oxaloacetate via citrate synthase) is transported into the cytoplasm by the tricarboxylate transport system:

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In the cytoplasm, citrate is cleaved by ATP-citrate lyase to yield cytosolic acetyl-CoA:

Oxaloacetate, one of the products of the citrate lyase reaction, is converted into pyruvate, which then returns to the mitochondria:

NADP+ molecules reduced in this reaction by malate dehydrogenase (malic enzyme) serve as reducing equivalents in the reductive reactions of fatty acid biosynthesis.

Acetyl-CoA produced in the citrate lyase reaction is carboxylated by cytosolic acetyl-CoA carboxylase to yield malonyl-CoA, the immediate precursor for fatty acid synthesis:

The biotin-containing enzyme acetyl-CoA carboxylase is a regulatory enzyme that controls The rate of the entire fatty acid biosynthesis process. Citrate acts as its allosteric modulator; thus, in this context, citrate performs two crucial Functions: serving as an acetyl-CoA carrier and as an activator of acetyl-CoA carboxylase.

The subsequent reactions of this biosynthetic pathway are carried out directly by a multienzyme system known as the fatty acid synthase system, or palmitate synthase. This enzyme complex, with a total molecular mass of ~ 400 kDa, integrates seven Enzymes involved in The biosynthesis of palmitic acid (Fig. 14.5). The central component of this system is the acyl carrier protein (ACP), which functions during synthesis as a molecular tether that sequentially transfers synthesis intermediates from one synthase enzyme to the next. The prosthetic group of ACP, 4'-phosphopantetheine (a derivative of pantothenic acid), forms thioester bonds with each intermediate product during the construction of the fatty acid carbon chain, analogously to the process occurring in the pyruvate dehydrogenase enzyme complex (see Ch. 12, Sec. 12.1.3).

Fig. 14.5. Diagram of the palmitate synthase enzyme complex:

I - acetyltransferase; II - malonyltransferase; III - 3-ketoacyl-ACP synthase;

IV - 3-ketoacyl-ACP reductase; V - 3-hydroxyacyl-ACP dehydratase; VI - enoyl-ACP reductase

The fatty acid synthase system catalyzes the overall reaction for constructing the 16-carbon chain of the palmitic acid molecule from one molecule of acetyl-CoA and seven molecules of malonyl-CoA:

In this synthesis, the acetyl-CoA molecule serves as a primer to which two-carbon units from malonyl-CoA are sequentially added. The elongation of the higher fatty acid carbon chain proceeds from its methyl and oxo groups (positions 16 and 15, respectively) toward the carboxyl group of palmitate.

Palmitate synthesis is initiated by acetyltransferase and malonyltransferase, which catalyze the attachment of the synthesis substrates—the acetyl and malonyl groups—to two SH-groups of the synthase. Subsequently, the elongation of the fatty acid proceeds cyclically in four stages (Fig. 14.6).

Fig. 14.6. Reaction sequence for a single cycle of palmitate synthesis

First, the acetyl group is transferred from the Cysteine SH group of the synthase to the malonyl group attached to the phosphopantetheine arm of ACP through the action of 3-ketoacyl-ACP synthase. In the Condensation reaction between these two groups, CO₂ is released, which was previously incorporated into the malonyl-CoA molecule during the acetyl-CoA carboxylase reaction.

Next, acetoacetyl-S-ACP is reduced by 3-ketoacyl-ACP reductase in the presence of NADPH to form 3-hydroxybutyryl-S-ACP. This is followed by a dehydration reaction catalyzed by 3-hydroxyacyl-ACP dehydratase, yielding trans-Δ²-butenoyl-S-ACP.

At The final stage of the first synthesis cycle, the double bond of trans-Δ²-butenoyl-S-ACP is reduced by enoyl-ACP reductase, and the butyryl group is transferred from butyryl-S-ACP to the free cysteine SH group of the synthase—occupying the site initially held by the acetyl group.

This is followed by a new cycle of reactions extending the carbon chain by another two-carbon unit. These cycles are repeated seven more times, ultimately resulting in the end product, palmitoyl-S-ACP. This completes the fatty acid synthesis process. Upon the action of a hydrolase, the palmitic acid (C₁₆) molecule is cleaved from the ACP synthase.

It should be noted that the synthesis of a single palmitate molecule requires the energy of seven high-energy ATP bonds to form thioester bonds in the acetyl-CoA and malonyl-CoA substrate molecules, as well as the reducing equivalent of 14 NADPH molecules for the saturation of double bonds.

NADPH molecules for the reductase reactions of synthesis come from two sources: glucose-6-phosphate oxidation reactions via the Pentose Phosphate Pathway and malate oxidation in the malate dehydrogenase reaction.

The fatty acid synthase system is incapable of synthesizing fatty acids with a carbon chain length exceeding 16 atoms. However, further elongation of the palmitic acid carbon chain is possible through The Endoplasmic reticulum enzyme systems by adding acetyl groups from malonyl-CoA, or in mitochondria, where acetyl groups from acetyl-CoA molecules are added.

Palmitic and stearic acids serve as precursors for two monounsaturated fatty acids: palmitoleic (C₁₆Δ⁹) and oleic (C₁₈Δ⁹) acids, which are formed via the action of acyl-CoA desaturase, molecular oxygen, and NADPH:

Animal Tissues (mammals) cannot synthesize linoleic (C₁₈Δ^{9, 12}) and α-linolenic (C₁₈Δ^{9, 12, 15}) acids, which are precursors for two other important polyunsaturated fatty acids—γ-linolenic (C₁₈Δ^{6, 9, 12}) and arachidonic (C₂₀Δ^{5, 8, 11, 14}) acids—essential precursors of biologically active compounds known as Eicosanoids.

The BIOSYNTHESIS OF HIGHER fatty acids in animal tissues is regulated by molecular mechanisms operating at both THE CELLULAR LEVEL and the Organism level as a whole (Fig. 14.7).

Fig. 14.7. Schematic diagram of The regulation of acetyl-CoA carboxylase activity

At the cellular level, mechanisms of Allosteric Regulation and covalent modification govern the Key Enzymes of long-chain fatty acid biosynthesis. The activity of the primary regulatory enzyme of this biosynthetic pathway—acetyl-CoA carboxylase, which determines the rate of fatty acid synthesis—is controlled by allosteric modulators: the enzyme is activated by citrate and inhibited by palmitoyl-CoA.

Palmitoyl-CoA also acts as an inhibitor of palmitoyl synthase via negative feedback, or retroinhibition.

During the absorptive period, the hormone Insulin activates a phosphatase that dephosphorylates inactive acetyl-CoA carboxylase, converting it into an active state. In the postabsorptive period, as well as during stress, Physical Exercise, and other states, Hormones such as Glucagon, adrenaline, and others acting through the adenylate cyclase system inactivate acetyl-CoA carboxylase by phosphorylating it via active protein kinase A.

Fatty acid synthesis is also regulated by diet. A carbohydrate-rich diet leads to the induction of A number of carbohydrate and Lipid METABOLISM enzymes that catalyze The conversion of glucose breakdown intermediates into fatty acids.

Malonyl-CoA, produced in the acetyl-CoA carboxylase reaction, also inhibits carnitine acyltransferase activity, thereby suppressing the uptake and Oxidation of Fatty acids in mitochondria. A decrease in cytosolic malonyl-CoA levels and the consequent activation of carnitine acyltransferase stimulate the translocation of fatty acids from the cytoplasm into mitochondria and their subsequent β-oxidation.