e key role of fatty acidn Metabolism in Type 2 Diabetes: A Tribute to J.

Denis McGarry e late Denis McGarry, a long-time to this book, wrote a scien article in 1992 entitled what if Minkowski had been ageus- An alternative angle on diabetes. Oskar Minkowski(1858-1931) I Josef von Mering were the firs to recognize that the pancreas duces a substance that regulates the level of blood glucose; their earch ultimately led to the isolation of insulin from cells of the icreas.he cocept of diabetes as a disease of carbohydrate metab- m began with the ancient observations that diabetic urine is sweet. Klore tells us that Dr. minkowski put his finger into the urine of a betic patient, tasted it, and noted that it was as sweet honey. To s day, diabetes is widely considered to be due to an imbalance in bohydrate metabolism. Dr.McGarry raised the humorous supposi-n that if minkowski had been ageustic (i.e.,he could not taste), but ild smell the acetone in the urine, diabetes would have been clas-ed as a disease of lipid rather than carbohydrate metabolism. This n important insight into type 2 (noninsulin-depended) diabetes, ich is associated with obesity and resistance to insulin. Because most of the human energy reserve isTriacylglycerols this fuel xtremely important particulary during fasting. After a meal, glucose apidly removed from the blood and either used for energy or stored glycogen; within hours humans switch to the oxidation of fatty acids the major source of energy, with ketone bodies becoming impor-t during prolonged fasting. After 3 days of fasting, even the brain tabolizes ketone bodies. The metabolism of fat over the limited glu- e supply is referred to as fuel sparing. Fatty acids play an important e in this process by inhibiting glucose utilization in skeletal muscle. 1963, Philip randle and colleagues proposed what is now known the randle hypothesis, which states that fatty acid oxidation blocks cose metabolism by inhibiting key regulatory enzymes involved in cose utilization. Including phosphofructokinase and pyruvate dehy- ogenase complex (see capter 15) and inhibits glycogen synthesis in letal muscle. Fatty acids also markedly reduce the uptake of glucose m the blood by blodcking the recruitment of GLUT 4 from the endo- smic reticulum to the activation specific isoforms of protein kinase C, as well as the transcription fac- NF-B, by acyl CoAs generated by fatty acid metabolism.

Ogen. In addition, Triacylglycerols are stored without associated water, whereas glycois hydrophilic and binds about twice its weight of water. Therefore, the energy yield gram of stored Triacylglycerol is about 4 times that of hydrate glycogen. Humans e much more fuel as Triacylglycerol than as glycogen. The average 70-kg human stores ut 250 g of glycogen in liver and muscle. This represens about 1000 kcal of energy, ch is less than one days energy requirement. By contrast, the same individual stores 5 kg of Triacylglycerol, which rovides sufficien energy to survive several week of vation. Unlike stores of glycogen and amino acids, which are limited, Triacylglycerol es, can greatly expand, depending on caloric intake. This makes Triacylglycerol an llent fuel store, but it can also lead to obesity and, in some cases, diabetes (clin. Rs. 17.1 and 17.2). adipose tissue is a significant source of hormones that regulate etite, and subcutaneous fat is also important in temperature regulation because it ides a layer of insulation.

17.3. interorgan transport of fatty acids and their rimary products

In all mammals, the transport and storage of fatty acids is regulated by dietary status. Triacylglycerol is stored in the fed state, with net deposition in adipose tissue. During fasting, Triacylglycerols in adipose tissues is hydrolyzed and the products are distributed throughout the bo be used for energy production. As fasting progresses, the liver converts fatty acids to the ke bodies, acetoacetate, and -hydroxybutyrate, which are released into the blood and bec a major source of energy for many tissues. These processes are summarized in figure 17.6 The transport of lipids presents unique problems because these molecules (especiall acylglycerols, cholesterol and its esters) are insoluble in water. Consequently, lipids are t ported through the bloodstream in plasma lipoproteins or bound to proteins. In addi the transport of Triacylglycerols bacross membranes usually involves breakdown to sm constituens by lipases.

Lipid Transport In The Fed State

Triacylglycerols in the diet are digested in the stomach and small intestine by gastri pancreatic lipases (p. 1052). The principal roducts are 2-monoacylglycerols and fatty acids, which are

absorbed by the epithelial calls that line the small intestine. T cells assemble the absorbed fatty acids and monoacylglycerols into Triacylglycerols (p. Ich are than packaget into chylomicrons, a Triacylglycerol-rich plasma lipoprotein 1051). Chylomicrons are secreted into lymph and then circulate in the bloodstream. The liver is another source of Triacylglycerols in the feed state. Fatty acids are synthesized in s tissue from excess carbohydrate and amino acids. These fatty acids are assembled into Triacylglycerols and packaged into very low-density lipoprotein (VBLDL), a second Triacylglycerol-n lipoprotein, which is secreted into the blood streem (p.724). The Triacylglycerols in chylomicrons and VLDL are hydrolyzed by lipoprotein lipase, Ated on the surface of capillary endothelial cells in tissues such as adipose tissue and skeletal iscle. The apoprotein apoC-II, which is found in chylomicrons and VLDL, activates the cess by binding the lipoproteins to the enzyme. The products of lipoprotein lipase action free fatty acids and glycerol. The free fatty acids are utilized in the tissue whare hydrolysis curs .The glycerol is transported through the bloodstream and taken up by the liver where s used in glycolysis or gluconeogenesis. Lipoprotein lipase is present at high levels in adipose tissue, cardiac muscle, and skeletal scle, allowing these tissues to utilize Triacylglycerols from lipoproteins. In adipose tissue, products of lipoprotein lipase are taken up and assembled intoTriacylglycerols, allowing deposition of fuel (p.688). in muscle, the fatty acid products of lipase action are taken up d used to generate energy, although some Triacylglycerol synthesis occurs in this tissue. Figure 17.6 interorgan transport of fatty acids, () in the fed state, there is net deposition of Triacylglycerol in adipose tissue. The sources are dietary fat and fatty acids synthesized in the liver from excess carbohydrate and amino acids. (b) in the fasted state, Triacylglycerols are hydrolyzed and free fatty acids and glycerol are released into the blood. *during fasting, the liver shyntesize ketone bodies, which become a major in the blood as fasting progresses Pid Transport In The Fasted State

tricylglycerols stored in adipose tissue are mobilized for use as fuel in the fasted state. Is process is initiated by hormone-sensitive lipase, which is located within adipocytes. Is enzyme

is activated when it is phosphorylated by cAMP-dependent protein kinase conversely, insulin inhibits the activity of this enzyme by inducing its dephosphoryla-n. the protein, perilipin, which coats the surface of fat droplets, is also important in s regulation. When perilipin is not phosphorylated,it blocks lipase acces to the Triacylglycerol. When perilipen is phosphorylated by protein kinase A, hormone-sensitive lipase nslocates to the surface of the fat droplet and hydrolyzes Triacylglycerols. The principal duct of this enzyme is monoacylglycerol and free fatty acids. Other enzymes complete hydrolysis. A summary of the regulation of Triacylglycerol metabolism is presented in ble 17.2. the regulation results in the net mobilization of fatty acids during fasting and deposition after a meal. The blace of Triacylglycerol synthesis and hydrolysis helps assuare adequate energy stores and avoid obesity (see clin. Cross 17.1 and 17.2). other ases, including adipose triglyceridle lipase, may also play a role in the degradation of tricylglycerols. These may contribute to slow unregulated lipolysis. Other lipases rapidly complete the hydrolysis, releasing fatty acids and glycerol into blood. The fatty acids are referred to as free fatty acids although they circulate bound serum albumin. Each albumin molecule can bin 10 fatty acids, so its binding capac- is very high. Free fatty acids bound to albumin, however, are a relatively small fraction the total lipid in plasma when one considers the presence of the lipids in the plasma oproteins (p.724). however, these free fatty acids turn over rapidly, so they represent a ificant fraction of the lipid flux through the blodstreem. The hydrolysis ofTriacylglycerols present in adipose tissue and in plasma lipoproteins o produces free glycerol. This glycerol is used by the liver. Which contain high levels glycerol kinase, an enzyme that synthesize glycerol 3-phosphate from glycerol and ATP (see figure 15.39, p. 625). Glycerol is metabolized sparingly in other tissues dueto low levels of this enzyme. Hepatic glycerol 3-phosphate dehydrogenase converts glycerol 3-phospha to dihydroxyacetone phosphate, which enters the glycolytic pathway in the fed state. In the fasted state, it is converted to glucose via gluconeogenesis. During prolonged fasting, whe much of the bodys energy is derived from stored fat, the glycerol produced by the hydroly of Triacylglycerol in adipose tissue is an important substarate for gluconeogenesis in the liver.

17.4. SYNTHESIS OF FATTY ACIDS; LOPONGENESIS

Glucose Is The Major Precursor For Fatty Acid Synthesis

Dietary carbohydrate in excess of that needed for energy production and glycog synthesis is converted to fatty acids in the liver during the fed state in mammals. Gluc provides the carbon for fatty acid synthesis (via acetyl CoA) as well as the reducing equi lents (NADPH) required for this process, other substrates, such as amino acids, can a contribute to lipogenesis.

Pathway Of Fatty Acids Biosynthesis

The synthesis of fatty acids occur in the cytosol using acetyl CoA produced from glucose from other precursor (i. e., the carbon skeletons of amino acids). The saturated C 16 acid, pa itic acid, is synthesized first, and all other fatty acids are made by its modification. Fatty a are synthesized by sequential adition of two-carbon units from acetyl CoA to the active carboxyl end of a growing chain by fatty acids synthase. In bacteria, fatty acids synthase complex of several proteins, whereas in mammalin cell it is a single multifunctional pro.

Formation of malonyl is the commitment step of fatty acid synthe

Synthesis of fatty acids from acetyl CoA requires the activated intermediate, malonyl c which is made by carboxylation of acetyl CoA by acetyl-CoA carboxilase (figure I ) the reaction requires ATP and bicarbonate as the source of CO2. In the first step c linked to abiotin moiety on the enzyme, using energy derived from ATP hydrolysis CO2 is then transferred to acetyl CoA. This reaction is similar to the carboxylation of p vate to oxaloacatate by pyruvate carboxylase (p. 622). Acetyl-CoA carboxylase catalize the rate-limiting step in fatty acid synthesis. This en existen in an inactive, protomeric state and, when activated, as linear polymers. The mammals enxyme is activated by citrate or isocitrate. This represents feed-forward activation of fatty synthesis because citrate is exported from the mitochondria to generate cytosolic acetyl for fatty acid synthesis (see the section on the citrate cleavage pathway p. 684). Acetyl carboxylase is also inhibited by long-chain acyl CoAs, resulting in fadback in hibition o acid synthesis by the end product of the pathway. Phosphorilation of acetyl-CoA carbo by both cyclic-AMP-dependen

protein kinase A and AMP-dependen protein kinase I this enzyme. The importance of this regulation is discussed on page 702. Reaction sequence for the synthesis of palmitic acyd

This firs step catalyzed in fatty acid synthesis is the transfer of the primer molecule, an acetyl or butyryl group from CoA to a4-phpsphopantetheine moiety on acetyl c protein (ACP), a protein constituent of the multyenzyme complex (figure 17.8, tion 1). The phosphopantetheine unit is identical with that in CoA; both are derived fro Figure 17.8 pathway of fatty acid synthesis.

Nin, pantothenic acid. In bacteria ACP is a small protein, whereas in mammals it is a ain of fatty acids synthase. Six or seven two-carbon units are added to the enzyme com- (depending on whether acetyl CoA or butyryl CoA is the primer) in a repetitive sequence actions until the palmitate molecule is completed. The reaction sequence, staring with cetyl CoA primer and leading to butyryl-ACP, is presented in figure 17.8. A round of fatty acids elongation is initiated by the addition of two carbon atoms to the n in a three-step process. Firs the fatty acids chain is transferred from the 4-phosphoetheine moiety of ACP to a cysteine sulfhydryl group of -ketoacyl-ACP synthase (reac2a). the SH group of ACP then accepts a malonyl unit from malonyl CoA (reaction) then the condensation step links the growing acyl chain to C2 of the malonyl group, the loss of CO2 (reaction 2c). this is the same CO2 that was added by acetyl-CoA oxylase, so the carbon atoms in palmitare are derived entirely from acetyl CoA. The product is -ketoacyl-ACP. This intermediate is reduced to hydroxyacylACP using OPH as electron donor (reaction 3). -hydroxyacyl-ACP is dehydrated to an enoyl- (reaction 4) and then reduced to a saturated acyl CoA, using a second molecule of OPH as reducent (reaction 5). The growing fatty acids proceeds through six more reac-cycles to produce palmitoyl-ACP, which then is acted on by a thioesterase to produce free itic acid (figure 17.9). the specificity of this enzyme determines the chain length of the acid product. Note that at this stage, the sulfhydryl groups of ACP and synthase are both so that another cycle of fatty acid synthesis can begin. The released product is converted lmitoyl CoA, preparing it for modification or incorporation into complex lipids.

Mammalian fatty acid synthase is a multifunctional polypeptide The reactions outlined above are the basic sequence for fatty acid synthesis in all organisms the enzyme complex, fatty acid synthase, catalyzes all these reactions, but its structure and properties very considerably. In Escherichia coli, the complex is composed of separate enzymes. By contrast, mammalian fatty acid synthase is composed of two identical sub-unites each of which is a miltienzyme polypeptide that contains all of the catalytic activites. There are also variations in the enzyme between mammalian speciesand tissues. The growing fatty acid chain is always bound to the multifunctional protein and is transferred sequentially between the 4-phosphopantetheine group of ACP, which is a domain of the protein, and the cysteine sulfhydryl group of -ketoacyl-ACP synthase, during the condensation reaction (reaction 2, figures 17.8(p.681) and 17.10). The levels of fatty acids synthase in tissues are controlled by the rate of its synthesis and degradation. As shown in table 17.3, insulin increases the rate of fatty acidsynthesis by stimulating transcription of the fatty acid synthase gene, thereby increasing enzyme levels

the citrate cleavage pathway provides acetyl CoA and NAPDH for lipogenesis in the cytosol glucose breakdown in the liver via glycolysis result in the production of pyruvate, which is converted to acetyl CoA in the mitokondria by pyruvate dehydrogenes complex. however, the syntesis of fatty acids is a cytosolic process and acetyl CoA is not readily transported across the inner mitochondrial membrane. the citrate cleavage pathway over comes this problem. citrate, formed by citrate synthase in the tricarboxylic acid cycle, transported across the mitochondrial inner membrane via the tricarboxylic transport (see figure 14.49,p.578). the citrate is then cleaved in the cytosol by ATP citrate lya ( also called citrate cleavage enzyme) to from acetyl CoA and oxaloacetate (figure 17.11) this reaction is not a reversal of citrate synthase, since it requires the hydrolysis of ATP. a mentioned previously, citrate has a second role in fatty acid synthesis as an allosteric activatour of acetyl-CoA carboxylase, the rate-limiting enzime in lipogenesis the oxaloacetate generated by citrate cleavage in the cytosol is not readily transport back into the mitochondria. it is instead reduced to malate by the cytosolic isoform NA malate

dehyidrogenese. the malate then undergoes oxidative decarboxylation to pyiruvate catalyzed by NADP malate dehydrogenese (also called malic enzime). the pyruvate enter the mitochondria for further metabolism. the removal of citrate from the mitochondria (cataplerosis) must be by its replacement (anaplerosis) in order maintain tricarbosilic acid cycle flux.this is achived by the conversion of pyruvate to oxaloacetate pyruvate carboxylase (p.622), the major anaplerotic enzyme in mitochondria.

the conversion of oxaloacetate to pyruvate by the citrate cleavage pathway transfer a pair of electron from NADH to generate NADPH that is used for fatty acid synthesis. the source of the NADH for this process in the glyceraldehyde-3-phosphate dehydrogena reaction in the

glycolytic pathway. the metabolism of glucose thus provides both the carbon atoms and the reducting equivalents required for lipogenesis

modification of fatty acids humans can synthesize the other fatty acids they require from palmitate. except for some polyunsaturated fatty acids (see table 17.1, p. 675). fatty acids are modified as the CoA derivatives by three processes: elongation, desaturation, and hydroxylation.

elongation reactions

fatty acids elongation in mammals occurs in either the endoplasmic reticulum or the mitochondria. these processes are slightly different in the two locations. in the endoplasmic reticulum, fatty acyl CoA are elongated by reactions similar to the ones catalyzed by cytosolic fatty acids synthase: malonyl CoA is the source of two carbon units and NADPH provides the reducing power. the prefered elongation substrate is palmitoyl CoA, which is converted almost exclusively to stearate (18:0) in most tissues. the brain, however, kontain one or more additional elongation systems, which synthesize longer-chain acids (up to C24) that are required for membrane lipids. these elongation systems also use malonyl CoA. fatty acid elongation in mitochondria uses acetyl CoA, and both NADH and NADPH as electron donors (figure 17.12). this systems operates by reversal of the pathway of fatty acid (lambang beta) -oxsidation (p.692) whit the exception that NADPH-linked enoyl- coA reductase (last step of elongation) replaces FAD- linked acyl-CoA dehydrogenase (first step in-b- oxidation) . this process has little actifity whit acyl CoA substrates of 16 Carbons or longer, it serves primarily to elongate shorter molecules.

formation of monoenoic Acids by stearoyl-CoA desaturase invertebrate, fatty acid desaturation occurs in the endoplasmic reticulum, and the reaction and enzymes that introduce cis double bond are significantly different from the acyl-CoA dehydrogeneses of mitochondrial B oxidation. desaturation is carried out by monooxygeneses, which have fatty acyl CoA, NADH and O2 as substrates (p.695). the three components of the system are the desaturase enzyme, cytochrome b5, and NADPH-cytochrome b5 reductase. the overall reaction is the initial step in the formation of unsaturated fatty is the formation of the A9 double bond by stearoyl-CoA desaturase in palmitic or stearic acid to produce palmitolic or oleic acid, respectively. the synthesis of unsaturated fatty acids is important for regulating the fluidity of triacylglycerols and membrane phospholipids. it is also required for the synthesis of cholesterol esters in the liver and waxy secretions in the skin,which preferentially use newly synthesized, rather than dietary fatty acid. expression of stearoyl-CoA desaturase is highly regulated by both dietary and hormonal mechanisms. insulin, triiodothyronine, hydrocortisone, and dietary

cholesterol incrase gene transcription and thus the levels of stearoyl-CoA desaturase in the liver, whereas dietary polyunsaturated fatty acids have the opposite effect.

formation and modification of polyunsaturated fatty acids polyunsaturated fatty acids, particularly arachidonic acid (20:4), are precursors of important signaling molecules; prostagladins, thromboxanes, and leukotrienes (p.737). polyunsaturated fatty acids are also required for the synthesis of complex lipids, particulary in the nervous system. these fatty acids can olso undergo nonenzymatic oxidation reactions, creating products that damage cellular constituens and may have pathological effects. in mammals, doubel bonds can be added only to the proximal half of fatty acyl CoAs; they cannot be added beyond C9. consequently, linolelic (18:3) are obtained in the diet from plants and cold-water fish. there are two series of polyunsaturated fatty acids. the distal doubel bond of linoleic acid is six carbons from the methyl group; it is referred to as n-6 or w-6. a second series, n-3 or w-3, has the distal doubel bond three carbons from the methyk group. one isomer of linolenic acid is part of the series

the esential fatty acids are modified by elongation and desaturation to from the polyunsaturated fatty acids found in mammals. a variety of polyunsaturated fattyb acidss, are synthesized by human using disaturases that introduce dobel bons at position 5, or 6 (figur 17:14). these enzymes act only in the synthesis of plyunsaturrated fatty acids because they can use only substrates with a double bond at position 9. this elongation and desaturation occurs in the second order. conversion of linoleic acid to arachidonic acid (all cis-5, 8, 11, 14-eicosatetraenoic acid) is an example of a rection sequence. formation of hydroxy fatty acids in nerve tissue two different processes produce a-hydroxy fatty acids in vertebrates. one occurs in the mitochondria of many tissues and acts on relatively short-chain fatty acids. the other has been demonstrated only in the nervous system, where it produces long-chain fatty acids hydroxylated

on C2 that are required for the formation of some myelin lipids. the enzyme in the brain that catalyzed this reaction is a monooxygenase that requires O2 dan NADH or NADPH and preferentially uses C22 and C24 fatty acids. this process is closely coordinated with the synthesis of sphingolipids that contain hydroxylated fatty acids (see p. 729)

fatty acid synthase can produce fatty acids other than palmitate

the principal fatty acids synthesis by humans for

energy storage are palmitate and its

modification products. however, smaller amounts of different fatty acids are synthesized for other purpose. two examples are the roduction of the fatty acids shorter than palmitate in the mammary gland and the synthesis of branched-chain fatty acids in some secretory glands. milk contains fatty acids with chains that are shorter than palmitate. the relative amounts of the fatty acids produced by the mammary gland vary with species dan with the physiological state of the animal. in ruminants, the pathway of fatty acids synthesis, which normally produces palmitate, is modified to synthesize fatty acids as short as C4. this is accomplished by the expression of soluble thioesterases, which cleave the shorter chains from fatty acids synthase. human milk contains no fatty acids with chains shorter than 10 carbons.

there are relatively few branched-chain fatty acids in vertebrates. until recently, their metabolism has been studied mostly in bacteria such as mycobacteria, where they are present in a greater variety and amount. simple, branch-chain fatty acids are synthesized by tissues of vertebrates for specific purpose, such as the production of waxes in sebaceous glands, and avian preen glands, and the elaboration of structures in echo-locating systems of porpoises.

most of the brached-chain fatty acids in vertebrates are mehylated darivatives of saturated, straight-chain acids that are synthesized by fatty acids synthase. when methylmalonyl CoA is used as a substrate instead of malonyl CoA, a methyl side chain is inserted into the fatty acid by the following reaction.

regular reduction steps then follow. these reactions occurs in many tissues at a rate several orders of magnitude lower than the utilization of malonyl CoA in fatty acid synthesis. the proportion of branched-chain fatty acids that are synthesized is largely governed by the relative availability of the two precursors. an increse in branching can occur by decreasing the ratio of malonyl CoA. to Methylmalonyl CoA. A malonyl-CoA decarboxylase that is responsible for this decrease occurs in many tessues. it is been suggested that an increased concentration of methylmalonyl CoA, which occurs in vitamin B12 deficiency, can leas to excessive production of branch-chain fatty acids triacylglycerols are synthesized in most tissues from fatty acyl CoAs and a glycerol precursor, glycerol 3-phosphate (figure 17.16). glycerol 3-phosphate is derived from several sources. in most tissues, it is synthesized by the reduction of dihydroxyacetone phosphate in the fed state, the dihydroxyacetone phosphate is derived from glucose via glycolysis: in the fasted state in adipose tissue and liver, glycerol 3-phosphate is derived from glyceroneogenesis (p.689). in the liver, there is an additional source of glycerol 3-phosphate; glycerol can be phosphorylated by glycerol kinase, which is very active in this tissue.

fatty acids are activated for fruther metabolism by conversion to their CoA esters in the following :

this two step reaction heas an acyl adenylate (fatty acyl-AMP) as an intermediate. the over all reaction is driven by the hydrolysis of the pyrophosphate product to two Pi.

the synthesis of triacylglycerols involves the formation of phosphatidic acid, which is formed by sequential acylations of glycerol 3-phosphate to from lysophospatidic and then phospatidic acid (Figure 17.17). Phosphatidic acid is used for triacylglycerol synthesis by hydrolysis of the phosphate group by phosphatidate phosphatase to yielow

1 7,6 UTILIZATION OF FATTY ACIDS FOR ENERGY PRODUCTION

Fatty acids in the circulation are taken up by cells and used for energy production, primarily in mitochondria, in a process integrated with energy generation from other sources. Fatty acids are broken down to acetyl CoA in the mitochondria, with the production of NADH

FADH2. These three products are then used in the mitochondrial matrix for energy production via the tricarboxylic acid cycle and oxidative phosphorylation. The utilization of fatty acids for energy production varies considerably from tissue to

tissue and depends to a significant extent on metabolic status, that is, fed or fasted, exercising or resting. Most tissues can use fatty acids as a fuel. Fatty acids are a major energy source in cardiac and skeletal muscle, but the brain does not oxidize fatty acids because they cannot cross the blood-brain barrier. Mammalian red blood cells cannot oxidize fatty acids because they lack mitochondria, the site of fatty acid oxidation. During fasting, the liver converts acetyl CoA, generated by fatty acid oxidation and the breakdown of amino acids, into ketone bodies, which become a major fuel after 2-3 days of fasting. Most tissues, including the brain, adapt to fasting by utilizing these ketone bodies.

p-Oxidation of Straight-Chain Fatty Acids Is a Major Energy-Producing Process CoA esters of fatty acids are the substrates for oxidation. For the most part, the pathway of fatty acid oxidation is similar, but not identical to, a reversal of the process of fatty acid synthesis. That is, two-carbon fragments are removed sequentially from the carboxyl end of the fatty acid by enzymatic oxidation, hydration, and oxidation to form a (3-keto acid, which is then split by thiolysis. The pathway is called /3-oxidation because carbon 3 (the (3-carbon) is oxidized twice prior to cleavage.

fatty acids are its activated by conversion fatty acyl CoA The first step in the utilization of a fatty acid is its activation to a fatty acyl CoA. This reaction is catalyzed by enzymes in the endoplasmic reticulum and outer mitochondrial membrane and is the same one used in the synthesis of triacylglycerols. The fatty acyl Coast are released into the cytosol.

Carnitine Carries Acyl Groups across the Inner Mitochondrial Membrane Most long chain fatty acyl CoAs are formed outside the mitochondria, but are oxidized in the mitochondrial matrix. The mitochondrial membrane is impermeable to CoA and its derivadyes. Fatty acids are transported into the mitochondria using carnitine (4-trimethylamino-3hydroxybutyrate) as a carrier. The process is outlined in Figure 17.20.

the acyl group is transferred from CoA to carnitine by carnitine palmitoyltransferase I (CPT I) on the outer mitochondrial membrane. Aryl carnitine and free carnitine are then exchanged across the inner mitochondrial membrane by carnitine-acylcarnitine translocase. Sally, the fatty aryl group is transferred back to CoA by carnitine palmitoyltransferase II CPT II) on the matrix side of the inner mitochondrial membrane. This process functions narily in the transport of fatty acyl CoAs with 12-18 carbons. Genetic abnormalities in system lead to serious pathological consequences (Clin. Corr. 17.4). By contrast, entry of f i t t er-chain fatty acids is independent of carnitine; they cross the inner mitochondrial memories as free fatty acids and become activated to their CoA derivatives in the matrix. CPT an important site for the regulation of fatty acid oxidation because its rate controls the entry of fatty acids into the mitochondria and therefore determines the supply of ubstrate /3-oxidation in the mitochondrial matrix. Malonyl CoA, which is the product of acetyl-Ci carboxylase and a key intermediate in fatty acid synthesis (p. 680), is an inhibitor of CPT -Oxidation Is a Sequence of Four Reactions -oxidation is a series of four reactions that act on a fatty aryl Co A to produce an acetyl C and a new acyl CoA that is two carbon atoms shorter than the initial substrate (Figure II) . Once a fatty aryl CoA is formed at the inner surface of the inner mitochondrial membr.,n it can be oxidized by acyl-CoA dehydrogenase, a flavoprotein that uses FAD as the elecr acceptor (Reaction 1). The product is an enoyl CoA with a trans double bond betwee i tl C2 and C3 atoms and enzyme-bound FADH2. As in the tricarboxylic acid cycle, the FAL)P transfers its electrons to enzymes of oxidative phosphorylation, regenerating FAD. The second step in (3-oxidation is hydration of the trans-double bond to a 3-L-hydto1 acyl CoA, which is oxidized to a 3-ketoacyl CoA intermediate, with the generation of NA in the third step. The final step is cleavage of the chain by ketothiolase, generating acetyl and a fatty-aryl CoA that has been shortened by two carbon atoms. This shortened aryl is ready for the next round of oxidation, starting with acylCoA dehydrogenase. In mossues, the acetyl CoA will be used by the tricarboxylic acid cycle and the FADH2 and N will be reoxidized by the oxidative phosphorylation pathway with the production of K Each of the four reactions shown in Figure 17.21 is catalyzed by several diff enzymes that have specificity for substrates of different chain lengths. For example, at four enz y mes catal y ze the first dehydrogenation step. These are very long-chain, long-c medium-chain, and short-chain acyl-CoA dehydrogenases (VLCAD, LCAD, MCAT)SCAD). VLCAD, which

oxidizes straight-chain acyl CoAs ranging from C12 to C2-+ ff ern rom t h e ot h er famil y members in that it is membrane associated. MCAD has chain length specificity but is most active with C6 and C8 substrates, whereas the order preference for SCAD is C4 > C6 > C8. LCAD is involved in initiating the oxidat

Triacyiglycerol/Fatty Acid Cycle The triacylglycerol that is stored in adipose tissue is hydrolyzed to free fatty acids (FFA) during fasting to provide energy for tissues such as skeletal and cardiac muscle, and indirectly to the brain via ketone bodies. Hormones, most notably insulin, control this process. As the level of insulin falls during fasting, the rate of triacylglycerol hydrolysis (lipolysis) increases, resulting in FFA release from adipose tissue.

A surprising aspect of this process is the fate of the FFA; in fasted humans up to 65% of this FFA is re-esterified to triacylglycerol in the liver and other peripheral tissues. In liver the triacylglycerol products are released into the blood as VLDL and sent back to the adipose tissue for deposition as triacylglycerol. This process has been termed the triacylglycerol/fatty acid cycle.

The synthesis of triacylglycerol in mammalian tissues requires glycerol 3-phosphate, hich can be derived from dietary glucose via glycolysis in the fed state. During fasting, when low insulin inhibits glucose utilization, the glycerol 3-phosphate for the re-etherification of FFA is generated by glyceroneogenesis, an abbreviated version of gluconeogenesis. In this pathway, pyruvate- or compounds that can generate pyruvate, such as alanine or lactate is converted to glycerol 3phosphate via dihydroxyacetone phosphate (Figure 17.19). Recent studies have shown that glyceroneogenesis and not glycolysis is the predominant pathway for the synthesis of glycerol 3-phosphate, even during the fed state. The key enzyme in the pathway of glyceroneogenesis is phosphoenolpyruvate carboxykinase (PEPCK), which is very active in brown and white adipose tissue. If the expression of the PEPCK gene is ablated in the adipose tissue of mice, glyceroneogenesis is inhibited and triacylglycerol storage is reduced. Conversely, overexpression of the PEPCK gene in a pose tissue of transgenic mice increases the rate of glyceroneogeneai resulting in obesity.

The metabolic logic of the triacylglycerol/fatty acid cycle most lily resides in the importance of fatty acids as a fuel during starvation. i order to ensure that there is sufficient FFA in the blood, more FFA i released from fat cells than is required; what is not used is restcrih, ill to triacylglycerol and redeposited in adipose tissue, at a small energy cost. The triacylglycerol/fatty acid cycle consumes 3%-6% of the energy in a molecule of triacylglycerol; it is apparently better to have needed fuel available and pay for it energetically, than to run sho a critical time! The rate of FFA reesterification in the triacylglyer fatty acid cycle is a key factor in determining the steady-state cone, tration of FFA in the blood, a parameter that is directly involved in the etiology of Type 2 diabetes (see Clin. Corr. 17.2, p. 677). '11, thiazolidinediones, a class of antidiabetic drugs, induce the activities PEPCK in adipose tissue and increase the rate of FFA resterificar, to triacylglycerol via glyceroneogenesis in this tissue, supporting important role of this process in maintaining lipid homeostasis. diacyiglycerol, which is then acylated to triacyiglycerol (Figure 17.18). Phosphatidic acid so a key intermediate in the synthesis of other glycerolipids (p. 712). In an alternative ;nway, dihydroxyacetone phosphate is acylated, reduced to lysophosphatidic acid, and acylated. a second time to form phosphatidic acid (see Figure 17.16). Although this is a major pathway of triacyiglycerol synthesis, it is important for the synthesis of memne lipids with ether-linked alky chains. Triacylglycerol synthesis follows a different pathway in epithelial cells of the small or stine. These cells take up 2-monoacylglycerols and free fatty acids from the gut, which are the major digestion products of dietary triacyiglycerols by pancreatic lipase. An enzyme in the mucosal cells acylates these monoacylglycerols using acyl CoAs as substrates. The resulting diacylglycerols can then be acylated to form triacyiglycerols, which are packaged into chylomicrons. Analysis of human triacyiglycerols shows that each position of glycerol is esterified nth fatty acids of distinct composition. Saturated fatty acids are found preferentially at position 1 and unsaturated fatty acids at positions 2 and 3. Two main factors that determine the fatty acid composition at each position on glycerol are the specificity of the ayltransferases involved and the relative availability of different fatty acids in the fatty aryl CoA pool.

Mobilization of Triacylglycerols Requires Hydrolysis The first step in mobilizing stored fatty acids for energy production is hydrolysis of treacle glycerol. Several lipases catalyze this reaction and the sequence of hydrolysis of the three acv] chains is determined by the specificities of the lipases involved. Treacle glycerol Synthesis Occurs during Fasting As Part of a Triacylglycerol-Fatty Acid Cycle Involving Glyceroneogenesis The release of free fatty acids from adipose tissue is a critical metabolic adaptation to fasting. The quantity of fatty acids released by adipose tissue, however, exceeds the amount used for energy by other tissues. As much as 60% of these fatty acids are redeposited in adipose tissue as triacylglycerols. Both liver and adipose tissue play a major role in this process (Clin. fCorr.17.3). In the fed state, the glycerol 3-phosphate for triacylglycerol synthesis is derived om glucose via glycolysis. During fasting, however, the entry of glucose into adipose tissue is limited because the insulin concentration is low and glucose is being used by other sues. In this dietary state, glycerol 3-phosphate is synthesized in both the adipose tissue and the liver by glyceroneogenesis. As shown in Figure 17.19, substrates that enter the carboxylic acid cycle, such as pyruvate, glutamate, or aspartate, can support net glycero-neogenesis. This pathway is essentially an abbreviated version of gluconeogenesis, in which Manlate formed in the tricarboxylic acid cycle leaves the mitochondria and is converted to oxaloacetate in the cytosol. Phosphoenolpyruvate carboxykinase then converts oxaloacetate O phosphoenolpyruvate, which is converted to dihydroxyacetone phoshate and then top Ivicerol 3-phosphate, which is used for triacylglycerol synthesis. The key enzyme in this Bess is phosphoenolpyruvate carboxykinase; its activity is induced in liver and adipose le during fasting.