Principles of Biochemistry Volume 2 - A. Lehninger 1985

Bioenergetics and Metabolism
Lipid Biosynthesis
The addition of each two-carbon unit proceeds in four stages

a. Condensation

In the first of the four stages of fatty acid carbon chain elongation, the acetyl and malonyl groups, covalently linked to the —SH groups of the synthase, undergo condensation to form an acetoacetyl group covalently bound to the —SH group of phosphopantetheine; simultaneously, a CO₂ molecule is released. This reaction is catalyzed by 3-ketoacyl-ACP synthase:

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Note that the acetyl group is transferred from the Cysteine —SH group to the malonyl group, which is attached to the phosphopantetheine —SH group; thus, it becomes the terminal two-carbon unit of the newly formed acetoacetyl group (Fig. 21-9). In this process, the acetyl group displaces the free carboxyl group of the malonate residue as CO₂. This reaction produces the exact same CO₂ that was originally incorporated into the malonyl-CoA molecule via the acetyl-CoA carboxylase reaction described above. Thus, during FATTY ACID Biosynthesis, Carbon dioxide is not used to build the covalent backbone of the fatty acid molecule; instead, it acts as a catalyst, since it is regenerated each time a new two-carbon unit is added by splitting off from the growing fatty acid chain.

Fig. 21-9. Condensation reaction in fatty acid biosynthesis. The decarboxylation of the malonyl group (shown in red) provides the driving force for The transfer of the acetyl group (on a gray Background) from Cys—SH to the 2nd carbon atom of the malonyl group. Upon completion of fatty acid synthesis, the initial acetyl group becomes the final two-carbon unit at the methyl end of the fatty acid molecule.

Why do Cells need to create a malonyl group from an acetyl group by adding CO₂, which is then split off again during The formation of acetoacetate? The answer to this question lies in the fact that when CO₂ is cleaved from the malonyl group, the reactivity of the remaining two-carbon fragment increases sharply, allowing it to rapidly react with the acetyl group (Fig. 21-9).

b. 3-Keto reduction

Acetoacetyl-S-ACP then undergoes reduction at the carbonyl group to yield D-3-hydroxybutyryl-S-ACP. In this reaction, catalyzed by 3-ketoacyl-ACP reductase, NADPH serves as the electron donor (Fig. 21.10).

Fig. 21-10. Three Sequential Stages of the fatty acid synthesis cycle. Upon completion of the final reaction, the butyryl group is transferred to the cysteine —SH group. Afterwards, the Fp—SH group is ready once again to bind the next incoming malonyl group of malonyl-CoA.

Note that the D-3-hydroxybutyryl group is not sterically identical to the L-3-hydroxyacyl intermediate formed during Fatty acid oxidation (Section 18.4,6).

c. Dehydration

During the Third Stage of the fatty acid synthesis cycle, D-3-hydroxybutyryl-S-ACP is dehydrated by 3-hydroxyacyl-ACP dehydratase to form trans-Δ²-butenoyl-S-ACP (Fig. 21-10).

d. Saturation

In the Fourth Stage, which completes one cycle of reactions carried out by the fatty acid synthase complex, the double bond of trans-Δ²-butenoyl-S-ACP is reduced, or saturated, by enoyl-ACP reductase to yield butyryl-S-ACP (Fig. 21-10). NADPH again acts as the electron donor in this reaction.

The butyryl group is subsequently transferred from the phosphopantetheine —SH group to the cysteine —SH group.

The newly formed, elongated acyl group now occupies the position at the —SH group to which the acetyl group was originally attached.

A new cycle of reactions leading to the elongation of the chain by another two-carbon unit begins with the transfer of the next malonyl group from malonyl-CoA to the phosphopantetheine —SH group of ACP (Fig. 21-11). Next, the butyryl group leaves the cysteine —SH group and displaces CO₂ in the malonyl group on HS—ACP. This results in a six-carbon acyl group covalently bound to the phosphopantetheine —SH group. During the next three Stages of the synthase cycle, the 3-keto group of the resulting acyl group is reduced, yielding a six-carbon saturated acyl group, much like in the previous reaction cycle. Then, the hexanoyl group is transferred from the phosphopantetheine —SH group to the cysteine —SH group.

After seven such cycles, the end product, palmitoyl—S-ACP, is formed. Chain growth terminates at the 16th carbon atom, after which the palmitic acid molecule is cleaved from the ACP molecule by the action of a hydrolytic enzyme.

Fig. 21-11. Initiation of the second turn of the fatty acid synthesis cycle. The butyryl group is linked to the —SH group of cysteine. The incoming malonyl group binds to the —SH group of Pp. At the condensation stage, the butyryl group attached to the Cys—SH group is exchanged for the free carboxyl group of the malonyl residue, which is thereby released as CO₂ (shown on a red background). In the resulting product—the six-carbon 3-ketoacyl group—four carbon atoms originate from malonyl-CoA, and the remaining two from the acetyl-CoA that initiated the reaction.

Note that the synthesis of palmitic acid requires two forms of chemical energy: the phosphate group energy of ATP and the reducing potential of NADPH. ATP is needed to form the thioester bond in acetyl-CoA and to build malonyl-CoA by adding CO₂ to acetyl-CoA, whereas NADP is used to reduce the double bonds.

The NADPH required for the reduction reactions in fatty acid biosynthesis comes from two different sources in various Cell types. In the Liver, NADPH is produced primarily via the Reactions of the Pentose Phosphate Pathway (Section 16.13), mainly driven by glucose-6-phosphate dehydrogenase.

In adipose cells (adipocytes), NADPH is generated predominantly through the action of malate dehydrogenase.

Both of these reactions leading to NADPH formation take place in the Cytosol, where the NADPH/NADP⁺ molar ratio is very high (approximately 75), providing the high reducing potential necessary for fatty acid synthesis. In contrast, the cytosolic NADH/NAD⁺ molar ratio is significantly lower (approximately 8 ∙ 10^-4). Thus, cytosolic NADPH is ideally suited to serve as the primary donor of hydrogen atoms utilized in reductive biosynthetic reactions.

Summarizing the above, the enzymatic biosynthesis of palmitic acid differs from its enzymatic oxidation in: 1) intracellular localization; 2) The Nature of the acyl carrier; 3) the form in which two-carbon units are added to or removed from the fatty acid chain; 4) the stereoconfiguration of the intermediate ß-hydroxyacyl compound; 5) the type of pyridine nucleotide utilized in reduction reactions; and 6) the involvement of CO₂. These differences (Table 21-1) demonstrate that these two opposing pathways are characterized by distinct Physical and Chemical parameters.

Table 21-1. Differences between the enzymatic biosynthesis and enzymatic oxidation of palmitic acid