Biological Chemistry - Berezov T. T., Korovkin B. F. 1998

Lipid Metabolism
Ketone Body Metabolism
Biosynthesis of Saturated Fatty Acids

Currently, The Mechanism of FATTY ACID Biosynthesis in Human and Animal organisms, as well as the enzyme systems catalyzing this process, has been studied in considerable detail. Fatty acid synthesis occurs in the Cell Cytoplasm, whereas Mitochondria are primarily responsible for the elongation of existing fatty acid chains. It has been established that palmitic acid (containing 16 carbon atoms) is synthesized in the cytoplasm of Liver Cells. In contrast, Fatty acids with 18, 20, and 22 carbon atoms are formed in the mitochondria of these cells from palmitic acid previously synthesized in the cytoplasm or from exogenous fatty acids derived from the intestine.

In other words, the mitochondrial system of fatty acid biosynthesis, which incorporates a slightly modified sequence of β-oxidation reactions, merely elongates medium-chain fatty acids already present in the Organism. Conversely, the complete de novo biosynthesis of palmitic acid from acetyl-CoA actively takes place in the Cytosol—that is, outside the mitochondria—via an entirely different pathway.

Extramitochondrial system of de novo fatty acid biosynthesis (Lipogenesis). This system is localized in the soluble (cytosolic) fraction of cells from various Organs, notably the liver, Kidneys, Brain, Lungs, mammary gland, and adipose tissue. Fatty acid biosynthesis proceeds with the participation of NADPH, ATP, Mn²⁺, and HCO₃^- (acting as a source of CO₂); acetyl-CoA serves as the substrate, and palmitic acid is the end product. The cofactor requirements for the processes of biosynthesis and β-Oxidation of Fatty acids differ significantly.

As noted, the building block for fatty acid synthesis in The Cell cytosol is acetyl-CoA, which is primarily supplied by the mitochondria. It has been demonstrated that citrate stimulates fatty acid synthesis in the cell cytosol. Furthermore, it is known that acetyl-CoA generated within mitochondria during The oxidative decarboxylation of Pyruvate and The oxidation of fatty acids cannot diffuse directly into the cell cytosol because the mitochondrial membrane is impermeable to this substrate. Consequently, intramitochondrial acetyl-CoA first reacts with oxaloacetate to yield citrate, a reaction catalyzed by the enzyme citrate synthase. The resulting citrate is transported across the mitochondrial membrane into the cytosol via a specialized tricarboxylate transport system.

In the cytosol, citrate reacts with HS-CoA and ATP, breaking down once again into acetyl-CoA and oxaloacetate—a reaction catalyzed by ATP-citrate lyase. Once in the cytosol, oxaloacetate is reduced to malate with the participation of cytosolic malate dehydrogenase. The malate is then returned to the mitochondrial matrix via a dicarboxylate transport system, where it is oxidized back to oxaloacetate, thereby completing the so-called shuttle cycle:

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There is yet another pathway for transferring intramitochondrial acetyl-CoA into the cell cytosol: via carnitine. As previously mentioned, carnitine acts as a carrier of acyl groups from the cytosol into the mitochondria during Fatty acid oxidation. Apparently, it can also perform this function in the reverse process—namely, The transport of acyl radicals, including the acetyl radical, from the mitochondria to the cell cytosol. However, when it comes to fatty acid synthesis, this transport pathway is not the primary one.

Formation of malonyl-CoA. The initial reaction in fatty acid biosynthesis is the carboxylation of acetyl-CoA, which requires bicarbonate, ATP, and manganese ions. This reaction is catalyzed by the enzyme acetyl-CoA carboxylase, which contains biotin as a prosthetic group. Avidin, a biotin inhibitor, suppresses this reaction as well as fatty acid synthesis as a whole.

It has been established that acetyl-CoA carboxylase consists of a variable number of identical subunits, each containing biotin, biotin carboxylase, a carboxyl-biotin-carrying protein, transcarboxylase, and a regulatory allosteric site; thus, it is a multienzyme complex.

The reaction proceeds in two stages: I — carboxylation of biotin involving ATP, and II — transfer of the carboxyl group to acetyl-CoA, resulting in The formation of malonyl-CoA:

Malonyl-CoA represents the first specific product of fatty acid biosynthesis. In the presence of the appropriate enzyme system, malonyl-CoA is rapidly converted into fatty acids.

The enzymatic systems responsible for fatty acid synthesis are called fatty acid synthetases. They are widely distributed in nature and can be isolated from various single-celled organisms, plants, and animal Tissues.

Fatty acid synthetases are divided into two groups. The first group comprises multienzyme, non-fractionable complexes with a Molecular Weight of approximately 500,000, in which all individual Enzymes are assembled into a compact Structure. Specifically, this group includes the fatty acid synthetases of animal tissues and Yeast.

The second group comprises fatty acid synthetases from which individual Enzymes can be isolated using protein Fractionation Methods. Such synthetases are found in A number of microorganisms (particularly *E. coli*) and plants. In other words, in these cases, all individual Enzymes of the synthetase system exist as autonomous Polypeptides.

The multienzyme complex known as fatty acid synthetase (or synthase) consists of six enzymes associated with the so-called acyl carrier protein (ACP). This protein is relatively thermostable, possesses two free HS groups (belonging to Cysteine and a phosphopantetheine residue attached to a Serine OH group), and participates in the synthesis of Higher Fatty Acids at virtually all stages. The molecular weight of ACP is approximately 10,000. In the synthetase system, this protein plays The Role of CoA. It should be noted that free ACP analogous to the microbial protein has not been detected in animal tissues. A multienzyme complex containing all the enzymes required for fatty acid synthesis has been isolated from the liver. The enzymes of this complex are bound so tightly to one another that all attempts to isolate them individually have been unsuccessful. Below is The sequence of reactions occurring during fatty acid synthesis:

The reaction cycle then repeats. Suppose the synthesis of palmitic acid (C₁₆) is underway. In this case, the formation of butyryl-ACP marks the completion of only the first of seven cycles, each of which begins with The addition of a malonyl-ACP molecule to the carboxyl end of the growing fatty acid chain. Concurrently, the distal carboxyl group of malonyl-ACP is released as CO₂. For instance, the butyryl-ACP formed in the first cycle reacts with malonyl-ACP:

Fatty acid synthesis is completed by the Cleavage of HS-ACP from acyl-ACP under the Influence of the enzyme deacylase. For example:

The overall equation for the synthesis of palmitic acid can be written as follows:

Alternatively, taking into account that the formation of one molecule of malonyl-CoA from acetyl-CoA consumes one molecule of ATP and one molecule of CO₂ (which is subsequently released), the overall equation can be presented in the following form:

The MAIN STAGES OF fatty acid biosynthesis can be represented as follows:

The General scheme of fatty acid synthesis in Escherichia coli is shown in Fig. 11.4. The sequence and Nature of the reactions involved in fatty acid synthesis—ranging from the formation of ß-ketoacyl-ACP (referred to as acetoacetyl-ACP in Fig. 11.4) to the completion of a single two-carbon chain elongation cycle—appear to be the reverse of fatty acid oxidation reactions. In reality, the pathways of fatty acid synthesis and oxidation do not overlap, not even partially. This becomes evident when considering certain distinct features of these two processes.

Compared to ß-Oxidation, fatty acid biosynthesis has several distinctive characteristics: fatty acid synthesis occurs primarily in the cell cytosol, whereas oxidation takes place in the mitochondria; the involvement of malonyl-CoA in biosynthesis, which is formed by the attachment of CO₂ to acetyl-CoA (in the presence of a biotin-dependent enzyme and ATP); the participation of the acyl carrier protein (HS-ACP) at all stages of synthesis; the Formation of the D(—)-isomer of the 3-hydroxy acid during biosynthesis, rather than the L(+)-isomer characteristic of fatty acid ß-oxidation; and the requirement for NADPH as a coenzyme in fatty acid synthesis. In the organism, NADPH is generated partly (about 50%) via the Pentose Phosphate Pathway and partly through other reactions, particularly the following:

Fig. 11.4. Synthesis of palmitic acid in Escherichia coli utilizing one molecule of acetyl-CoA and 7 molecules of malonyl-CoA. The first cycle of synthesis—the formation of butyryl-ACP—is shown in detail. The remaining 6 cycles are analogous to the first.

Formation of Unsaturated fatty acids. Fatty acid elongation.

Unlike plant tissues, animal tissues have a very limited ability to convert saturated fatty acids into unsaturated ones.

It has been established that the two most common monounsaturated fatty acids—palmitoleic and oleic acids—are synthesized from palmitic and stearic acids, respectively.

These transformations take place in the microsomes of liver and adipose tissue cells and require molecular oxygen, a reduced pyridine nucleotide system, and cytochrome b₅. Only the activated forms of palmitic and stearic acids undergo this conversion. The enzymes catalyzing these reactions are known as desaturases.

Alongside fatty acid desaturation (the Introduction of double bonds), chain elongation also occurs in the microsomes, and both processes can be combined and repeated. The elongation of a fatty acid chain proceeds through the sequential addition of two-carbon units to the corresponding acyl-CoA, utilizing malonyl-CoA and NADPH. The enzymatic system catalyzing fatty acid elongation is referred to as elongase. The scheme illustrates the pathways of palmitic acid conversion via desaturation and elongation reactions.