Oxidation of Odd-Chain Fatty Acids Requires Three Extra Reactions

Although most naturally occurring lipids contain fatty acids with an even number of carbon atoms, fatty acids with an odd number of carbons are found in significant amounts in the lipids of many plants and some marine organisms. Small quantities of the three-carbon propionate (CH₃-CH₂-COO^-) are added as a mold inhibitor to some breads and cereals, and thus propionate enters the human diet. Moreover, cattle and other ruminant animals form large amounts of propionate during fermentation of carbohydrates in the rumen. The propionate so formed is absorbed into the blood and oxidized by the liver and other tissues.
Long-chain odd-carbon fatty acids are oxidized by the same pathway as the even-carbon acids, beginning at the carboxyl end of the chain. However, the substrate for the last pass through the β-oxidation sequence is a fatty acyl-CoA in which the fatty acid has five carbon atoms. When this is oxidized and ultimately cleaved, the products are acetyl-CoA and propionyl-CoA. The acetyl-CoA is of course oxidized via the citric acid cycle, but propionyl-CoA takes a rather unusual enzymatic pathway, involving three enzymes. Propionyl-CoA is carboxylated to form the n stereoisomer of methylmalonyl-CoA (Fig. 16-12) by propionyl-CoA carboxylase, which contains the cofactor biotin. In this enzymatic reaction, as in the pyruvate carboxylase reaction (see Fig. 15-13), CO₂ (or its hydrated ion, HCO₃^- ) is activated by attachment to biotin before its transfer to the propionate moiety. The formation of the carboxybiotin intermediate requires energy, which is provided by the cleavage of ATP to ADP and Pi.
The D-methylmalonyl-CoA thus formed is enzymatically epimerized to its L stereoisomer by the action of methylmalonyl-CoA epimerase (Fig. 16-12). The L-methylmalonyl-CoA undergoes an intramolecular rearrangement to form succinyl-CoA, which can enter the citric acid cycle. This rearrangement is catalyzed by methylmalonylCoA mutase, which requires as its coenzyme deoxyadenosylcobalamin, or coenzyme B₁₂, derived from vitamin B₁₂ (cobalamin)

Fatty Acid Oxidation Is Tightly Regulated

In the liver, fatty acyl-CoAs formed in the cytosol have two major pathways open to them: (1) β oxidation by enzymes in the mitochondria or (2) conversion into triacylglycerols and phospholipids by enzymes in the cytosol. The pathway taken depends upon the rate of transfer of long-chain fatty acyl-CoAs into the mitochondria. The three-step process by which fatty acyl groups are carried from cytosolic fatty acylCoA into the mitochondrial matrix (Fig. 16-6) is rate-limiting for fatty acid oxidation. Once fatty acyl groups have entered the mitochondria, they are committed to oxidation to acetyl-CoA.
Malonyl-CoA, the first intermediate in the cytosolic biosynthesis of long-chain fatty acids from acetyl-CoA (Chapter 20), increases in concentration whenever the animal is well supplied with carbohydrate; excess glucose that cannot be oxidized or stored as glycogen is converted in the cytosol into fatty acids for storage as triacylglycerol. The inhibition of carnitine acyltransferase I by malonyl-CoA assures that the oxidation of fatty acids is inhibited whenever the liver is amply supplied with glucose as fuel and is actively making triacylglycerols from excess glucose.
Two of the enzymes of β oxidation are also regulated by metabolites that signal energy sufficiency. When the [NADH]/[NAD⁺] ratio is high, β-hydroxyacyl-CoA dehydrogenase is inhibited; in addition, high concentrations of acetyl-CoA inhibit thiolase.

Peroxisomes Also Carry Out β Oxidation

Although the major site of fatty acid oxidation in animal cells is the mitochondrial matrix, other compartments in certain cells also contain enzymes capable of oxidizing fatty acids to acetyl-CoA, by a pathway similar to, but not identical with, that in mitochondria. Peroxisomes are membrane-enclosed cellular compartments (see p. 38) in animals and plants, where hydrogen peroxide is produced by fatty acid oxidation and then destroyed enzymatically. As in the oxidation of fatty acids in mitochondria, the intermediates are coenzyme A derivatives, and the process consists of four steps (Fig. 16-13): (1) dehydrogenation; (2) addition of water to the resulting double bond; (3) oxidation of the β-hydroxyacyl-CoA to a ketone, and (4) thiolytic cleavage by coenzyme A. The difference between the peroxisomal and mitochondrial pathways is in the first step. In peroxisomes, the flavoprotein dehydrogenase that introduces the double bond passes electrons directly to O₂, producing H₂O₂ (Fig. 16-13). This strong and potentially damaging oxidant is immediately cleaved to H₂O and O₂ by catalase. By contrast, in mitochondria the electrons removed in the first oxidation step pass through the respiratory chain to O₂, and H₂O is the product, a process accompanied by ATP synthesis. In peroxisomes, the energy released in the first oxidative step of fatty acid breakdown is dissipated as heat.
High concentrations of fats in the diet result in increased synthesis of the enzymes of peroxisomal β oxidation in mammalian liver. Liver peroxisomes do not contain the enzymes of the citric acid cycle and cannot catalyze the oxidation of acetyl-CoA to CO₂. Instead, the acetate produced by fatty acid oxidation is exported from peroxisomes. Presumably some of this acetate enters mitochondria and is oxidized there.

Plant Peroxisomes and Glyoxysomes Use Acetyl-CoA from β Oxidation as a Biosynthetic Precursor

Fatty acid oxidation in plants occurs in the peroxisomes of leaf tissue and the glyoxysomes of germinating seeds. Plant peroxisomes and glyoxysomes are similar in structure and function. Glyoxysomes occur only during seed germination, and may be considered specialized peroxisomes. In plants, the biological role of β oxidation in peroxisomes and glyoxysomes is clear: it provides biosynthetic precursors from stored lipids. The β-oxidation pathway is not an important source of metabolic energy in plants; in fact, plant mitochondria do not contain the enzymes of β oxidation. During germination, triacylglycerols stored in seeds are converted into glucose and a wide variety of essential metabolites (Fig. 16-14). Fatty acids released from triacylglycerols are activated to their coenzyme A derivatives and oxidized in glyoxysomes by the same four-step process that occurs in peroxisomes (Fig. 16-13). The acetyl-CoA produced is converted via the glyoxylate cycle (Chapter 15) to four-carbon precursors for gluconeogenesis (Chapter 19). Glyoxysomes, like peroxisomes, contain high concentrations of catalase, which converts the H2O2 produced by β oxidation to H2O and O2. The β-Oxidation Enzymes Have Diverged during Evolution Although the β-oxidation reaction sequence in mitochondria is essentially the same as that in peroxisomes and glyoxysomes, the mitochondrial enzymes differ significantly from their isozymes in those compartments. These differences apparently reflect an evolutionary divergence that occurred very early, with the separation of gram-positive and gram-negative bacteria (see Fig. 2-6). In mitochondria, each of the four enzymes of β oxidation is a separate, soluble protein, similar in structure to the analogous enzyme in gram-positive bacteria. In contrast, the enzymes of peroxisomes and glyoxysomes are part of a complex of proteins, at least one of which contains two enzymatic activities in a single polypeptide chaih (Fig. 16-15). Enoyl-CoA hydratase and L-β-hydroxyacyl-CoA dehydrogenase activities both reside in a single, monomeric protein (Mr = 150,000), closely similar to the bifunctional protein of the gram-negative bacterium E. coli. The evolutionary selective value of retaining both types of β-oxidation system in the same organism is not yet apparent.