Fatty Acid Metabolism PDF

# Fatty Acid Metabolism ## Introduction In addition to carbohydrates, other molecules may serve as sources of energy for ATP production or as end products of anabolic pathways. Lipids in particular can store significant amounts of energy. Consequently, lipid catabolism serves as a source of energy...

# Fatty Acid Metabolism ## Introduction In addition to carbohydrates, other molecules may serve as sources of energy for ATP production or as end products of anabolic pathways. Lipids in particular can store significant amounts of energy. Consequently, lipid catabolism serves as a source of energy in many cells and provides the energy necessary to carry out gluconeogenesis in the liver. When cells have sufficient ATP, they may convert acetyl-CoA molecules to fatty acids and triglycerides to store energy for later use. This lesson describes the catabolism and anabolism of lipids, with a focus on fatty acids, and explains how lipid metabolism is regulated. ## 13.1.01 Lipid Transport and Catabolism Lipids are generally hydrophobic and poorly soluble in the aqueous environments found in most parts of the body. Therefore, lipid transport through the bloodstream requires the assistance of amphiphilic molecules (ie, molecules with both hydrophilic and hydrophobic groups) such as transport proteins and phospholipids. The hydrophobic portions of these transport molecules interact favorably with hydrophobic lipids, while the hydrophilic portions interact favorably with the aqueous environment. After a lipid-rich meal, lipids are transported using molecules called lipoproteins, which consist of the components shown in Figure 13.1. - Apolipoprotein - Triacylglycerol - Unesterified cholesterol - Esterified cholesterol - Phospholipid monolayer - Lipoprotein **Figure 13.1 General structure of a lipoprotein** To facilitate transport through the bloodstream and lymphatic system, dietary lipids in the intestine are first emulsified by bile salts (see Biology Chapter 15), which allows them to be accessed by intestinal hydrolytic enzymes. The hydrolyzed products can then be absorbed by intestinal cells, where free fatty acids are converted to triacylglycerides (also called triglycerides or triacylglycerols). The triacylglycerides derived from dietary sources are then packaged into lipoproteins called chylomicrons, which also contain some cholesterol and cholesteryl esters. Lipoproteins are surrounded by a phospholipid monolayer, which helps to solubilize the lipids for transport to the tissues. Triglycerides stored in the liver may similarly be packaged into another type of lipoprotein called very-low-density lipoproteins (VLDLs), which can then be transported to other tissues. Once chylomicrons and VLDLs deposit their triglycerides in the target tissues (eg, muscle, adipose), they become chylomicron remnants and intermediate-density lipoproteins (IDLs), respectively, which are enriched in cholesterol and cholesteryl esters compared to their precursors. IDLs and chylomicron remnants commonly return to the liver, where they deposit cholesterol and any remaining triglycerides. IDLs may also be metabolized into low-density lipoproteins (LDLs), which are even more enriched in cholesterol and serve to deliver cholesterol to various peripheral tissues as needed. High-density lipoproteins (HDLs) may assist in delivering cholesterol from peripheral tissues back to the liver by sequestering excess cholesterol that has been excreted by the peripheral tissues. The pathways of various types of lipoproteins throughout the body are shown in Figure 13.2. **Figure 13.2 Overview of pathways for different types of lipoproteins** Once triglycerides and fatty acids are delivered to their target tissues, they may be further metabolized for energy or for the formation of glycerophospholipids, as needed. During the further metabolism of a triglyceride, each of the ester bonds may be hydrolyzed by lipase enzymes. When all three ester linkages are hydrolyzed, the result is three free fatty acids and one glycerol molecule. The fatty acids may then be digested by $β$-oxidation, as described in Concept 13.1.02. The glycerol molecule can be phosphorylated to become glycerol 3-phosphate, which can then be oxidized to form dihydroxyacetone phosphate (DHAP). Figure 13.3 shows degradation of a triglyceride into its components, along with the metabolic fates of those components. **Figure 13.3 Triglyceride degradation** ## 13.1.02 Fatty Acid Oxidation Fatty acid oxidation is, as the name suggests, the oxidation of fatty acids to form acetyl-CoA. Most commonly, fatty acids are digested by $β$-oxidation in the mitochondria, although $β$-oxidation can also occur in peroxisomes. $β$-Oxidation is so named because the oxidation occurs at the $β$-carbon (ie, the carbon that is two units away from the carboxylic acid group). Before $β$-oxidation can begin, fatty acids must enter the mitochondrial matrix. Short-chain fatty acids (those with fewer than six carbons) and some medium-chain fatty acids (which have six to 12 carbons) can enter the mitochondrial matrix directly. Within the matrix, these fatty acids react with coenzyme A to become fatty acyl-CoA. $β$-Oxidation can then proceed. In contrast, fatty acids with more than eight carbons typically require assistance to enter the matrix, and this assistance is largely provided by the carnitine shuttle shown in Figure 13.4. **Figure 13.4 The carnitine shuttle** Fatty acids that require this shuttle are first converted to acyl-CoA molecules. Note that an acyl-CoA molecule is any hydrocarbon chain attached to coenzyme A through a thioester linkage and should not be confused with acetyl-CoA, which is specifically a two-carbon chain with a thioester linkage to coenzyme A. As shown in Step 1 of Figure 13.4, acyl-CoA molecules form in the cytosol when fatty acids react with coenzyme A, as catalyzed by acyl-CoA synthetase (ACS). This reaction also requires hydrolysis of ATP to AMP and pyrophosphate (PP₁). Acyl-CoA freely passes across the outer mitochondrial membrane. The inner leaflet of this membrane (ie, the side that faces the intermembrane space) contains the enzyme carnitine acyl transferase I (CAT I), which catalyzes the transfer of the acyl group from coenzyme A to carnitine (Step 2 in Figure 13.4). This enzyme is also sometimes called carnitine palmitoyl transferase I (CPT I). The resulting molecule is acylcarnitine, which is transported into the mitochondrial matrix through a translocase (Step 3 in Figure 13.4). At the same time, free carnitine exits the mitochondria through the same translocase (ie, through antiport). Within the matrix, acyl carnitine transferase II (CAT II, also called carnitine palmitoyl transferase II, CPT II) catalyzes the transfer of the acyl group back onto coenzyme A, reforming acyl-CoA and free carnitine (Step 4 in Figure 13.4). The free carnitine can then exit the matrix through the antiporter to react with another acyl-CoA in the intermembrane space. Once an acyl-CoA molecule is in the mitochondrial matrix, it can undergo $β$-oxidation. This process consists of four repeating steps: oxidation, hydration, another oxidation, and deacetylation. 1. Oxidation by acyl-CoA dehydrogenase. Three different isoforms of this enzyme act on short-, medium-, and long-chain fatty acids, respectively. The reaction forms a trans double bond between the $α$-carbon and the $β$-carbon (ie, carbons 2 and 3) of the chain. This converts the acyl-CoA into a trans-enoyl-CoA as shown in Figure 13.5. The oxidation of these carbons is accompanied by the reduction of FAD to FADH2, which is a prosthetic group in the enzyme. The enzyme then transfers electrons from FADH2 to ubiquinone, forming ubiquinol and regenerating FAD. In this way acyl-CoA dehydrogenase fills a similar role to Complex II of the electron transport chain, and the resulting ubiquinol interacts with Complex III. **Figure 13.5 Oxidation of acyl-CoA by formation of a double bond between carbons 2 and 3 (the $α$- and $β$-carbons, respectively). FADH2 forms in the process.** 2. Hydration by enoyl-CoA hydratase. Enoyl-CoA hydratase is a lyase that adds water across the double bond of trans-enoyl-CoA molecules. This results in a hydroxyl group on carbon 3 (the $β$-carbon) of the chain, converting trans-enoyl-CoA to L-3-hydroxyacyl-CoA, also called L-B-hydroxyacyl-CoA. Figure 13.6 shows this reaction. **Figure 13.6 Hydration of trans-enoyl-CoA by enoyl-CoA hydratase.** 3. Oxidation by $β$-hydroxyacyl-CoA dehydrogenase. In this reaction, the hydroxyl group that was added in Step 2 is oxidized to a ketone, forming a 3-ketoacyl-CoA, also called a $β$-ketoacyl-CoA, as shown in Figure 13.7. Simultaneously, NAD+ is reduced to NADH. The NADH then enters the electron transport chain at Complex I. **Figure 13.7 Oxidation of L-B-hydroxyacyl-CoA to B-ketoacyl-CoA using B-hydroxyacyl-CoA dehydrogenase. NADH forms as well.** 4. Deacetylation by acetyl-CoA acyltransferase. In this reaction, coenzyme A attacks the $β$-carbon. The result is release of acetyl-CoA and a new acyl-CoA chain that is two carbons shorter than the original chain, as shown in Figure 13.8. Acetyl-CoA acyltransferase enzymes are also known as thiolase enzymes. **Figure 13.8 Splitting of B-ketoacyl-CoA into a shortened acyl-CoA and acetyl-CoA by acyl-CoA acyltransferase.** The new acyl-CoA chain can then repeat Steps 1-4, shortening by an additional two carbons with each round. Once a four-carbon chain is produced, it undergoes $β$-oxidation one more time and is split into two acetyl-CoA molecules. Therefore, a 16-carbon saturated fatty acid undergoes seven rounds of $β$-oxidation to produce eight acetyl-CoA molecules along with seven FADH2 and seven NADH molecules. Each acetyl-CoA molecule can enter the citric acid cycle. **Concept Check 13.1** How many rounds of $β$-oxidation can the following acyl-CoA molecule undergo? How many acetyl-CoA, NADH, and FADH2 molecules are produced if it undergoes all rounds? **Solution** ## Unsaturated Fatty Acids Unsaturated fatty acids have at least one C=C double bond in the chain. If the double bond is between an odd-numbered carbon and a higher even-numbered carbon (eg, between carbons 9 and 10), $β$-oxidation proceeds normally until the chain has been shortened enough that the double bond is between carbons 3 and 4. At this point, instead of forming a new double bond, the enzyme enoyl-CoA isomerase simply alters the position of the pre-existing double bond. In addition to moving the position of the double bond, enoyl-CoA isomerase also converts it from a cis double bond between carbons 3 and 4 to a trans double bond between carbons 2 and 3. The formation of a double bond between carbons 2 and 3 is the first step in $β$-oxidation of a saturated fatty acid and produces FADH2. However, in an unsaturated fatty acid the double bond needs only to have its position and configuration shifted. No oxidation-reduction reaction is required to produce the double bond because the bond is already present. Consequently, one fewer FADH2 molecule is produced during $β$-oxidation of a fatty acid of this form, and therefore 1.5 fewer ATP molecules are produced on average per fatty acid oxidized (Figure 13.9). **Figure 13.9 $β$-Oxidation of a fatty acid with a C=C double bond between an odd-numbered carbon and a higher even-numbered carbon.** In contrast, if the original fatty acid contains a double bond between an even-numbered carbon and a higher odd-numbered carbon (eg, between carbons 8 and 9), another mechanism must occur. In this case when the fatty acid is shortened to the point at which the double bond is between carbons 4 and 5, acyl-CoA dehydrogenase acts normally to introduce a double bond between carbons 2 and 3. However, the resulting fatty acid, which now has a pair of conjugated double bonds, is no longer able to fit into the active site of enoyl-CoA hydratase. Consequently, a different enzyme called 2,4-dienoyl-CoA reductase removes both double bonds and creates a new one between carbons 3 and 4 (Figure 13.10). This process consumes NADPH to yield NADP+. Enoyl-CoA isomerase can then act on the new double bond to place it between carbons 2 and 3, and $β$-oxidation again proceeds normally. **Figure 13.10 $β$-Oxidation of an unsaturated fatty acid with a double bond starting at an even-numbered carbon.** The consumption of NADPH in this case is energetically equivalent to consumption of NADH. This is because the enzyme nicotinamide nucleotide transhydrogenase interconverts these molecules in the mitochondria. Therefore, fatty acids of this nature effectively produce one fewer NADH than unsaturated fatty acids of the same length. Polyunsaturated fatty acids may contain double bonds of both types, and accordingly may produce fewer NADH molecules, fewer FADH2 molecules, or both (relative to a saturated fatty acid), upon $β$-oxidation. ## Odd-Chain Fatty Acids Fatty acids with an odd number of carbons undergo normal $β$-oxidation until the last round. In fatty acids with an even number of carbons, the final round of $β$-oxidation involves a four-carbon molecule (butyryl-CoA) that becomes two acetyl-CoA molecules. When the number of carbons is odd, however, the final round releases a three-carbon molecule called propionyl-CoA as shown in Figure 13.11. **Figure 13.11 Example of $β$-oxidation of an odd-chain fatty acid yielding propionyl-CoA.** Propionyl-CoA is not a substrate for $β$-oxidation. Instead, this molecule becomes carboxylated by combining with bicarbonate ($HCO3$ - ) to form D-methylmalonyl-CoA. This reaction is catalyzed by propionyl-CoA carboxylase and requires hydrolysis of ATP to ADP and P₁. D-Methylmalonyl-CoA is converted to L-methylmalonyl-CoA by methylmalonyl-CoA epimerase, and L-methylmalonyl-CoA is then converted to succinyl-CoA by methylmalonyl-CoA mutase. This process is shown in Figure 13.12. **Figure 13.12 Conversion of propionyl-CoA to succinyl-CoA.** The details of this pathway are unlikely to be tested on the exam, but it is important to note the overall conversion of propionyl-CoA to succinyl-CoA, and the comparison to even-chain fatty acids, which only produce acetyl-CoA. Although the carbons of acetyl-CoA can be incorporated into oxaloacetate through the citric acid cycle, they must first react with oxaloacetate at the beginning of the citric acid cycle to do so. Therefore, no net oxaloacetate is produced or consumed by this pathway. Consequently, from a net carbon standpoint, acetyl-CoA cannot be converted to glucose. In contrast, the propionyl-CoA produced by odd-chain fatty acids can be used for gluconeogenesis because succinyl-CoA is converted to oxaloacetate without first reacting with oxaloacetate. **Concept Check 13.2** Which of the following molecules produces more acetyl-CoA upon $β$-oxidation? Which produces more FADH2? **Solution** ## 13.1.03 Ketone Bodies In the liver, $β$-oxidation is largely used to provide the energy needed for gluconeogenesis. This is accomplished as the NADH and FADH2 from $β$-oxidation enter the electron transport chain and as the acetyl-CoA from $β$-oxidation enters the citric acid cycle to produce additional NADH, FADH2, and GTP. The purpose of gluconeogenesis in the liver is to provide glucose that can be exported to other tissues, which can then use that glucose during fasting. Often, $β$-oxidation produces more acetyl-CoA than is needed to power gluconeogenesis. Rather than using the excess acetyl-CoA in the citric acid cycle within liver cells, acetyl-CoA itself can be converted to other forms that can be exported into the bloodstream and sent to other tissues. However, as discussed previously, acetyl-CoA cannot be converted to glucose. Instead, it is converted to a class of molecules known as ketone bodies, as shown in Figure 13.13. **Figure 13.13 Ketone body synthesis from acetyl-CoA.** Ketone body synthesis begins when two acetyl-CoA molecules condense to form acetoacetyl-CoA as catalyzed by the final thiolase enzyme of $β$-oxidation (also known as acetyl-CoA acetyltransferase). Acetoacetyl-CoA then condenses with another acetyl-CoA molecule to form HMG-CoA as catalyzed by HMG-CoA synthase. HMG-CoA is then separated by HMG-CoA Iyase, forming another acetyl-CoA molecule and the first ketone body, acetoacetate. Acetoacetate can be exported directly, or it may be reduced to form D-$β$-hydroxybutyrate. Reduction of acetoacetate requires oxidation of NADH to NAD+. Note that although D-$β$-hydroxybutyrate does not contain a ketone functional group, it is still classified as a ketone body because it is derived from a ketone-containing molecule. A small percentage of acetoacetate is decarboxylated to form acetone, which does not generally provide energy to cells and is instead exhaled. High levels of acetone, which may be detected on the breath, are indicative of inability to digest glucose (and therefore a need to metabolize ketone bodies instead), due to either lack of nutrition or metabolic disorders such as diabetes. Acetoacetate and D-$β$-hydroxybutyrate are transported to muscles, including the heart, as well as other tissues, as shown in Figure 13.14. In these tissues, D-$β$-hydroxybutyrate is oxidized back to acetoacetate, producing NADH in the process. Therefore, transport of D-$β$-hydroxybutyrate from the liver to other tissues effectively also transports NADH from the liver to those tissues. **Figure 13.14 Export of $β$-hydroxybutyrate and acetoacetate from the liver to skeletal muscle.** Acetoacetate is converted to acetoacetyl-CoA and can then split into two molecules of acetyl-CoA via the final step of the $β$-oxidation pathway (ie, thiolase, also known as acetyl-CoA acetyltransferase). The rest of the steps are not needed to process acetoacetyl-CoA; because it is already in the fully oxidized state (ie, the $β$-carbon is already a ketone), no additional oxidation is needed and no NADH or FADH2 is generated in this process. The resulting two acetyl-CoA molecules can enter the citric acid cycle in the target tissue, providing energy. ## 13.1.04 Fatty Acid Synthesis Fatty acid oxidation primarily takes place when glycogen stores are depleted and energy is needed for gluconeogenesis. In other words, fatty acid catabolism occurs mostly during fasting. In contrast, when glucose is abundant (ie, in the well-fed state), excess acetyl-CoA from glycolysis and pyruvate decarboxylation may be stored as fatty acids. Acetyl-CoA synthesis occurs in the mitochondria. In contrast, fatty acid synthesis occurs in the cytosol. Consequently, acetyl-CoA must be transported out of the mitochondria when fatty acid synthesis is active. However, acetyl-CoA cannot freely cross the inner mitochondrial membrane. Instead, acetyl-CoA in the mitochondrial matrix exits through the citrate shuttle shown in Figure 13.15. **Figure 13.15 The citrate shuttle.** The first step of the citrate shuttle is a reaction between acetyl-CoA and oxaloacetate to form citrate (ie, the first step of the citric acid cycle). Transport proteins allow citrate to exit the mitochondrial matrix (Step 2 in Figure 13.15). The outer mitochondrial membrane is porous and also allows citrate to cross it to enter the cytosol. Cytosolic citrate in the well-fed state can then serve both as a substrate source for fatty acid synthesis and as an allosteric signal to feedback inhibit glycolysis (see Chapter 11). To act as a substrate for fatty acid synthesis, cytosolic citrate is split into acetyl-CoA and oxaloacetate by the enzyme citrate lyase (Step 3 in Figure 13.15). Recall that in the citric acid cycle, the condensation of acetyl-CoA and oxaloacetate is irreversible. Because of this irreversibility, citrate synthase cannot simply catalyze the reverse reaction, and instead citrate lyase couples the reaction to ATP hydrolysis. Therefore, transport of each acetyl-CoA into the cytosol requires consumption of one ATP molecule. As shown in Step 4 of Figure 13.15, oxaloacetate is then converted to malate by cytosolic malate dehydrogenase (the same enzyme used in the malate-aspartate shuttle). Various pathways allow malate to reenter the mitochondrial matrix, including direct entry through a transporter; a common pathway during fatty acid synthesis is decarboxylation to form pyruvate. This step involves malic enzyme, which couples the oxidation of malate with the reduction of NADP+, as shown in Step 5. NADPH is required for subsequent steps in fatty acid synthesis, so using this method provides needed material. Pyruvate enters the mitochondria through the same transporter that it uses to enter the citric acid cycle (Step 6 in Figure 13.15). During well-fed states, mitochondrial pyruvate may be carboxylated by pyruvate carboxylase (the same enzyme used in gluconeogenesis). This regenerates oxaloacetate, as shown in Step 7, which can then react with more acetyl-CoA. Carboxylation of pyruvate requires hydrolysis of another ATP molecule, bringing the energy cost per acetyl-CoA transported to two ATP equivalents when this pathway is used. ## Fatty Acid Synthesis Mechanism Fatty acid synthesis is facilitated by a complex enzyme known as fatty acid synthase. The acyl carrier protein (ACP) is a domain on the enzyme to which thioesters may be transferred. The active site of this domain includes a prosthetic group called pantothenic acid, which contains a thiol (-SH) group. In the first step of fatty acid synthesis, shown in Figure 13.16, the acetyl group of acetyl-CoA is transferred to the ACP thiol group, releasing free coenzyme A. The acetyl group is then transferred to the $β$-ketoacyl synthase (KS) domain by reacting with a cysteine side chain. At this point, the enzyme is primed for fatty acid synthesis. **Figure 13.16 Initial step of the fatty acid synthase mechanism that links acetyl-CoA to the KS domain.** The ACP domain continues to react with thioesters, but now an activated form of acetyl-CoA is required. The necessary molecule is called malonyl-CoA and forms by carboxylation of acetyl-CoA. Malonyl-CoA synthesis is catalyzed by acetyl-CoA carboxylase, which serves as a major rate-determining enzyme of both fatty acid synthesis and oxidation (see Concept 13.1.05). Once malonyl-CoA is formed, it reacts with the ACP domain and releases its coenzyme A group as shown in Figure 13.17. **Figure 13.17 Linkage of malonyl-CoA to the ACP domain of fatty acid synthase.** At this point, a repeating series of four steps is used to synthesize fatty acids. These steps are similar to those of $β$-oxidation in reverse: condensation, reduction, dehydration, and another reduction. 1. Condensation of malonyl and acetyl groups. In this step, the acetyl group on the KS domain is transferred to the middle carbon of the malonyl group attached to the ACP domain. This process, shown in Figure 13.18, releases CO2 and forms a four-carbon group that contains both a thioester and a ketone, with the ketone at the $β$ position. **Figure 13.18 A reaction between the acetyl and malonyl groups yields a four-carbon chain and releases CO2** 2. Reduction of the ketone group. The ketone at the $β$ position is reduced to an alcohol, while NADPH is oxidized to NADP+. This reaction is shown in Figure 13.19. The four-carbon unit remains bound to the ACP domain prosthetic group throughout this process. **Figure 13.19 Reduction of the ketone group to an alcohol with oxidation of NADPH to NADP+.** 3. Dehydration. The hydroxyl component of the alcohol is removed along with a hydrogen atom from the adjacent $α$ carbon, forming water. A double bond also forms between the two affected carbon atoms, resulting in an alkenyl group, as shown in Figure 13.20. **Figure 13.20 Dehydration of the alcohol to form an alkenyl group.** 4. Reduction of the alkene. The double bond that formed between two carbons in Step 3 is reduced to a single bond (ie, an alkyl group), while NADPH is oxidized to NADP+. The result is a saturated fatty acyl chain linked to the ACP domain (Figure 13.21). **Figure 13.21 Reduction of the alkenyl group to an alkyl group, along with oxidation of NADPH.** After the four steps in the ACP domain are complete, the new four-carbon fatty acyl group is transferred back to the KS domain, and a new malonyl-CoA molecule reacts with the ACP prosthetic group. Steps 1-4 are then repeated, except instead of transferring a two-carbon acetyl group to malonyl-CoA, the four-carbon fatty acyl group is transferred to yield a six-carbon chain (Figure 13.22). **Figure 13.22 Elongation of a fatty acyl chain by repeating rounds of reactions with malonyl groups and subsequent reductions.** Once the four ACP steps are completed again, the new six-carbon acyl chain is transferred to KS, and the process is repeated to form an eight-carbon chain, a 10-carbon chain, and so on. Typically, this repeats until a 16-carbon chain is produced. The 16-carbon chain is hydrolyzed from fatty acid synthase to yield palmitate. Subsequent reactions in the smooth endoplasmic reticulum may further modify the chain, such as as by extending it or by adding double bonds. Each fatty acid chain may then be esterified onto glycerol backbone for formation of a triglyceride for storage or for formation of a glycerophospholipid. **Concept Check 13.3** Glycolysis of how many glucose molecules is required to provide the carbon to synthesize one palmitate molecule? **Solution** ## Energy Cost of Fatty Acid Synthesis Each malonyl-CoA molecule requires one ATP equivalent to produce. Seven malonyl-CoA molecules are added to one acetyl-CoA molecule to make palmitate; therefore, seven ATP equivalents are required for this process. In addition, each round of four ACP steps requires two NADPH molecules (ie, 14 NADPH molecules are needed to produce palmitate). The net equation for palmitate synthesis is 8 acetyl CoA + 7 ATP + 14 NADPH + 14 H+ palmitate + 8 CoA + 7 ADP + 7 P₁ + 14 NADP+ + 6 H2O Note that the citrate shuttle also requires two ATP equivalents per acetyl-CoA transported when pyruvate is used for shuttling back into the mitochondria, and therefore this system requires an additional 16 ATP equivalents to transport the eight acetyl-CoA molecules. The overall cost of palmitate synthesis is shown in Figure 13.23. **Figure 13.23 Energy cost of palmitate synthesis including the cost of acetyl-CoA transport out of the mitochondrial matrix.** As mentioned previously, the citrate shuttle provides one NADPH per acetyl-CoA transported, which can then be used in fatty acid synthesis. However, two NADPH molecules are required for each round of the ACP mechanism. Therefore, the citrate shuttle does not provide enough NADPH to power fatty acid synthesis by itself. The primary source of cytosolic NADPH in most eukaryotic cells is the oxidative phase of the pentose phosphate pathway (see Lesson 11.3). In summary, fatty acid synthesis has a high energy cost. The benefit of this process is that fatty acids can store a large amount of chemical energy in a relatively small volume, and this energy can be used when the metabolic needs of the body are not being met by the diet (ie, during fasting). ## 13.1.05 Regulation of Fatty Acid Metabolism Fatty acid oxidation and fatty acid synthesis are opposed metabolic processes. Assembling acetyl-CoA molecules into fatty acids costs more energy than is obtained by oxidizing fatty acids to acetyl-CoA. Therefore, allowing both fatty acid synthesis and oxidation to occur simultaneously wastes energy. To avoid this waste, the two processes are reciprocally regulated, meaning when one process is active the other is inactive. One of the major factors that aids in regulation of fatty acid metabolism is compartmentalization. Fatty acid synthesis takes place in the cytosol, whereas $β$-oxidation occurs in the mitochondrial matrix. Acetyl-CoA cannot freely cross the inner mitochondrial membrane and must use shuttles such as the citrate shuttle. Similarly, many fatty acids require the carnitine shuttle to enter the mitochondria. Figure 13.24 shows the compartmentalization of $β$-oxidation and fatty acid synthesis. **Figure 13.24 $β$-Oxidation and fatty acid synthesis occur in different compartments within the cell.** Therefore, control over these processes involves regulation of both the enzymes involved and in the shuttles involved. This control is also governed in part by blood glucose levels. Under high-glucose conditions (eg, after a meal), the pancreas releases insulin into the bloodstream. When insulin binds to its receptors, a cascade is triggered that dephosphorylates acetyl-CoA carboxylase (ACC), the enzyme that produces malonyl-CoA for fatty acid synthesis. Dephosphorylation activates this enzyme and induces malonyl-CoA production, which upregulates the synthesis pathway overall (see Figure 13.25). In addition to its role as a substrate in fatty acid synthesis, malonyl-CoA inhibits carnitine acyltransferase I and slows the carnitine shuttle. This prevents many fatty acids from entering the mitochondria, which prevents them from undergoing $β$-oxidation. In this way, ACC activation both stimulates fatty acid synthesis and inhibits fatty acid oxidation, preventing a futile cycle. **Figure 13.25 In high-glucose conditions, insulin causes activation of acetyl-CoA carboxylase (ACC), which produces malonyl-CoA to upregulate fatty acid synthesis and downregulate $β$-oxidation.** Conversely, when blood glucose is low (eg, after a period of fasting), the pancreas releases glucagon, which induces the reverse effect of insulin. Glucagon binding to its receptor results in activation of AMP-activated protein kinase (AMPK), which phosphorylates ACC and inactivates it. This inactivation results in lower malonyl-CoA levels, which in turn causes less fatty acid synthesis. Simultaneously, carnitine acyltransferase I inhibition ceases as malonyl-CoA is depleted, allowing fatty acids to enter the mitochondria. By these means glucagon upregulates $β$-oxidation and downregulates fatty acid synthesis, as shown in Figure 13.26. **Figure 13.26 Upregulation of $β$-oxidation and downregulation of fatty acid synthesis under low blood glucose conditions**