24.4 How Is Cholesterol Synthesized? 753 that the thiolase reaction is more or less reversible but slightly favors the cleavage re- action. In the cholesterol synthesis pathway, subsequent reactions, including HMG- CoA reductase and the following kinase reactions, pull the thiolase-catalyzed con- densation forward. However, in the case of fatty acid synthesis, a succession of eight thiolase condensations would be distinctly unfavorable from an energetic perspec- tive. Given the necessity of repeated reactions in fatty acid synthesis, it makes better energetic sense to use a reaction that is favorable in the desired direction. Squalene Is Synthesized from Mevalonate The biosynthesis of squalene involves conversion of mevalonate to two key 5-carbon intermediates, isopentenyl pyrophosphate and dimethylallyl pyrophosphate, which join to yield farnesyl pyrophosphate and then squalene. A series of four reactions converts mevalonate to isopentenyl pyrophosphate and then to dimethylallyl pyrophosphate (Figure 24.34). The first three steps each consume an ATP, two for the purpose of forming a pyrophosphate at the 5-position and the third to drive the decarboxylation CRITICAL DEVELOPMENTS IN BIOCHEMISTRY The Long Search for the Route of Cholesterol Biosynthesis colleagues at Merck Sharpe & Dohme isolated mevalonic acid and also showed that mevalonate was the precursor of isoprene units. The search for the remaining details (described in the text) made the biosynthesis of cholesterol one of the most enduring and challenging bioorganic problems of the 1940s, 1950s, and 1960s. Even today, several of the enzyme mechanisms remain poorly understood. C H CH 3 CCH 2 CH 2 OH ( (a) (b) Isoprene Squalene Lanosterol Many steps) Cholesterol ᮡ (a) An isoprene unit and a scheme for head-to-tail linking of isoprene units. (b) The cycliza- tion of squalene to form lanosterol, as proposed by Bloch and Woodward. Heilbron, Kamm, and Owens suggested as early as 1926 that squa- lene is a precursor of cholesterol. That same year, H. J. Channon demonstrated that animals fed squalene from shark oil produced more cholesterol in their tissues. Bloch and Rittenberg showed in the 1940s that a significant amount of the carbon in the tetracyclic moiety and in the aliphatic side chain of cholesterol was derived from acetate. In 1934, Sir Robert Robinson suggested a scheme for the cyclization of squalene to form cholesterol before the biosyn- thetic link between acetate and squalene was understood. Squalene is actually a polymer of isoprene units, and Bonner and Arreguin suggested in 1949 that three acetate units could join to form five- carbon isoprene units (see figure a). In 1952, Konrad Bloch and Robert Langdon showed conclu- sively that labeled squalene is synthesized rapidly from labeled ace- tate and also that cholesterol is derived from squalene. Langdon, a graduate student of Bloch’s, performed the critical experiments in Bloch’s laboratory at the University of Chicago while Bloch spent the summer in Bermuda attempting to demonstrate that radio- actively labeled squalene would be converted to cholesterol in shark livers. As Bloch himself admitted, “All I was able to learn was that sharks of manageable length are very difficult to catch and their oily livers impossible to slice” (Bloch, 1987). In 1953, Bloch, together with the eminent organic chemist R. B. Woodward, proposed a new scheme (see figure b) for the cy- clization of squalene. (Together with Fyodor Lynen, Bloch re- ceived the Nobel Prize in Medicine or Physiology in 1964 for his work.) The picture was nearly complete, but one crucial question remained: How could isoprene be the intermediate in the trans- formation of acetate into squalene? In 1956, Karl Folkers and his 754 Chapter 24 Lipid Biosynthesis HMG-CoA reductase kinase (inactive) P HMG-CoA reductase kinase kinase HPO 4 2 – HMG-CoA reductase kinase phosphatase HMG-CoA reductase kinase (active) HPO 4 2 – HMG- reductase phosphatase P HMG-CoA reductase (inactive) HMG-CoA reductase (active) ATP ADP ATP ADP H 2 O H 2 O CoA FIGURE 24.33 HMG-CoA reductase activity is modulated by a cycle of phosphorylation and dephosphorylation. CH 2 OH H 3 C C OH – OOC – OOC CH 2 O H 3 C C OH CH 2 O CH 2 H 3 C C H 2 C C C H CH 2 O H 3 C H 3 C P ++ P P P P P CH 2 CH 2 CH 2 CH 2 P P P P P CH 2 C C H H 3 C H 3 C CH 2 CH 2 C C H CH 2 C C H CH 2 O P P P P + + 2 H 3 C H 3 C ATP + H 2 O ADP CO 2 ATP ADP ATP ADP NADP + NADPH H + Mevalonate Mevalonate kinase 5-Pyrophosphomevalonate Phosphomevalonate kinase Pyrophosphomevalonate decarboxylase Isopentenyl pyrophosphate Isopentenyl pyrophosphate isomerase Dimethylallyl pyrophosphate Isopentenyl pyrophosphate Isopentenyl pyrophosphate Farnesyl pyrophosphate Squalene FIGURE 24.34 The conversion of mevalonate to squalene. In the last step, two farnesyl-PP condense to form squalene. 24.4 How Is Cholesterol Synthesized? 755 and double bond formation in the third step. Pyrophosphomevalonate decarboxylase phosphorylates the 3-hydroxyl group, and this is followed by trans elimination of the phosphate and carboxyl groups to form the double bond in isopentenyl pyro- phosphate. Isomerization of the double bond yields the dimethylallyl pyrophosphate. Condensation of these two 5-carbon intermediates produces geranyl pyrophosphate; ad- dition of another 5-carbon isopentenyl group gives farnesyl pyrophosphate. Both steps in the production of farnesyl pyrophosphate occur with release of pyrophosphate, hy- drolysis of which drives these reactions forward. Note too that the linkage of isoprene units to form farnesyl pyrophosphate occurs in a head-to-tail fashion. This is the gen- eral rule in biosynthesis of molecules involving isoprene linkages. The next step—the joining of two farnesyl pyrophosphates to produce squalene—is a “tail-to-tail” con- densation and represents an important exception to the general rule. HUMAN BIOCHEMISTRY Statins Lower Serum Cholesterol Levels Chemists and biochemists have long sought a means of reducing serum cholesterol levels to reduce the risk of heart attack and car- diovascular disease. Because HMG-CoA reductase is the rate- limiting step in cholesterol biosynthesis, this enzyme is a likely drug target. Mevinolin, also known as lovastatin (see accompanying fig- ure), was isolated from a strain of Aspergillus terreus and developed at Merck Sharpe & Dohme for this purpose. It is now a widely pre- scribed cholesterol-lowering drug. Dramatic reductions of serum cholesterol are observed at dosages of 20 to 80 mg per day. Lovastatin is administered as an inactive lactone. After oral in- gestion, it is hydrolyzed to the active mevinolinic acid, a competitive inhibitor of the reductase with a K I of 0.6 nM. Mevinolinic acid is thought to behave as a transition-state analog (see Chapter 14) of the tetrahedral intermediate formed in the HMG-CoA reductase re- action (see figure). Derivatives of lovastatin have been found to be even more potent in cholesterol-lowering trials. Synvinolin lowers serum cholesterol levels at much lower dosages than lovastatin. Lipitor, shown bound at the active site of HMG-CoA reductase, is the most-prescribed drug in the United States, with annual sales of $9 billion. O R CH 3 H CH 3 O O O CH 3 (a) CH 3 R H O O HO 1 R=H Mevinolin (Lovastatin, MEVACOR ® ) 2 R=CH 3 Synvinolin (Simvastatin, ZOCOR ® ) CH 3 H CH 3 O CH 3 CH 3 H OH HO COO – H OH HO COO – Mevinolinic acid OH HO COO – H 3 C Mevalonate HO COO – H 3 C H OH SCoA Tetrahedral intermediate in HMG-CoA reductase mechanism CH CH 3 CH 2 F CH 3 NHC N Lipitor ® (Atorvastatin) (b) (c) (d) HMG-CoA reductase with NADP + (magenta), HMG (blue), and CoA (green) (pdb id = 1DQA) Lipitor ® (red) bound at the active site of HMG-CoA reductase (pdb id = 1HWK) ᮡ The structures of (a) (inactive) lovastatin, (active) mevinolinic acid, mevalonate, and (b) Lipitor (atorvastatin). (c) HMG-CoA reductase with NADP ϩ , HMG, and CoA. (d) Lipitor bound at the HMG-CoA reductase active site. 756 Chapter 24 Lipid Biosynthesis HO O H 3 CCH 3 H 3 C H 3 C H + H + CH 3 HO H 3 C H 3 C HO H 3 C H 3 C HO H 3 C H 3 C CR O H 3 C H 3 C OCR O CoASH SCoA Squalene Lanosterol Squalene monooxygenase Squalene-2,3-epoxide 2,3-Oxidosqualene: lanosterol cyclase 7-Dehydrocholesterol Many steps (alternative route) Cholesterol Cholesterol esters Desmosterol Acyl-CoA cholesterol acyltransferase (ACAT) Many steps FIGURE 24.35 Cholesterol is synthesized from squalene via lanosterol.The primary route from lanosterol involves 20 steps, the last of which converts 7-dehydrocholesterol to cholesterol. An alternative route produces desmosterol as the penultimate intermediate. 24.5 How Are Lipids Transported Throughout the Body? 757 Squalene monooxygenase, an enzyme bound to the ER, converts squalene to squalene-2,3-epoxide (Figure 24.35). This reaction employs FAD and NADPH as coenzymes and requires O 2 as well as a cytosolic protein called soluble protein activator. A second ER membrane enzyme, 2,3-oxidosqualene lanosterol cyclase, catalyzes the second reaction, which involves a succession of 1,2 shifts of hydride ions and methyl groups. Conversion of Lanosterol to Cholesterol Requires 20 Additional Steps Although lanosterol may appear similar to cholesterol in structure, another 20 steps are required to convert lanosterol to cholesterol (Figure 24.35). The enzymes responsible for this are all associated with the ER. The primary pathway involves 7-dehydrocholesterol as the penultimate intermediate. An alternative pathway, also composed of many steps, produces the intermediate desmosterol. Reduction of the double bond at C-24 yields cholesterol. Cholesterol esters—a principal form of cir- culating cholesterol—are synthesized by acyl-CoAϺcholesterol acyltransferases (ACAT) on the cytoplasmic face of the ER. 24.5 How Are Lipids Transported Throughout the Body? When most lipids circulate in the body, they do so in the form of lipoprotein complexes. Simple, unesterified fatty acids are merely bound to serum albumin and other proteins in blood plasma, but phospholipids, triacylglycerols, cholesterol, and cholesterol esters are all transported in the form of lipoproteins. At various sites in the body, lipoproteins interact with specific receptors and enzymes that transfer or modify their lipid cargoes. It is now customary to classify lipoproteins according to their densities (Table 24.1). The densities are related to the relative amounts of lipid and protein in the complexes. Because most proteins have densities of about 1.3 to 1.4 g/mL, and lipid aggregates usually possess densities of about 0.8 g/mL, the more protein and the less lipid in a complex, the denser the lipoprotein. Thus, there are high-density lipoproteins (HDLs), low-density lipoproteins (LDLs), intermediate- density lipoproteins (IDLs), very-low-density lipoproteins (VLDLs), and also chylomicrons. Chylomicrons have the lowest protein-to-lipid ratio and thus are the lowest-density lipoproteins. They are also the largest. Lipoprotein Complexes Transport Triacylglycerols and Cholesterol Esters HDL and VLDL are assembled primarily in the ER of the liver (with smaller amounts produced in the intestine), whereas chylomicrons form in the intestine. LDL is not synthesized directly but rather is made from VLDL. LDL appears to be the major cir- culatory complex for cholesterol and cholesterol esters. The primary task of chy- lomicrons is to transport triacylglycerols. Despite all this, it is extremely important to note that each of these lipoprotein classes contains some of each type of lipid. The relative amounts of HDL and LDL are important in the disposition of cholesterol in the body and in the development of arterial plaques (Figure 24.36). The structures of Composition (% dry weight) Lipoprotein Density Diameter Class (g/mL) (nm) Protein Cholesterol Phospholipid Triacylglycerol HDL 1.063–1.21 5–15 33 30 29 8 LDL 1.019–1.063 18–28 25 50 21 4 IDL 1.006–1.019 25–50 18 29 22 31 VLDL 0.95–1.006 30–80 10 22 18 50 Chylomicrons Ͻ0.95 100–500 1–2 8 7 84 Adapted from Brown, M., and Goldstein, J., 1987. In Braunwald, E., et al., eds., Harrison’s Principles of Internal Medicine, 11th ed. New York: McGraw-Hill; and Vance, D., and Vance, J.,eds., 1985. Biochemistry of Lipids and Membranes. Menlo Park, CA: Benjamin/Cummings. TABLE 24.1 Composition and Properties of Human Lipoproteins FIGURE 24.36 Photograph of an arterial plaque.The view is into the artery (orange), with the plaque shown in yellow at the back. Science Photo Library/Photo Researchers, Inc. 758 Chapter 24 Lipid Biosynthesis the various lipoproteins are approximately similar, and they consist of a core of mo- bile triacylglycerols or cholesterol esters surrounded by a single layer of phospholipid, into which is inserted a mixture of cholesterol and proteins (Figure 24.37). Note that the phospholipids are oriented with their polar head groups facing outward to inter- act with solvent water and that the phospholipids thus shield the hydrophobic lipids inside from the solvent water outside. The proteins also function as recognition sites for the various lipoprotein receptors throughout the body. A number of different apoproteins have been identified in lipoproteins (Table 24.2), and others may exist as well. The apoproteins have an abundance of hydrophobic amino acid residues, as is appropriate for interactions with lipids. A cholesterol ester transfer protein also as- sociates with lipoproteins. Lipoproteins in Circulation Are Progressively Degraded by Lipoprotein Lipase The livers and intestines of animals are the primary sources of circulating lipids. Chy- lomicrons carry triacylglycerol and cholesterol esters from the intestines to other tis- sues, and VLDLs carry lipid from liver, as shown in Figure 24.38. At various target sites, particularly in the capillaries of muscle and adipose cells, these particles are de- graded by lipoprotein lipase, which hydrolyzes triacylglycerols. Lipase action causes progressive loss of triacylglycerol (and apoprotein) and makes the lipoproteins smaller. This process gradually converts VLDL particles to IDL and then LDL parti- cles, which are either returned to the liver for reprocessing or redirected to adipose tissues and adrenal glands. Every 24 hours, nearly half of all circulating LDL is re- moved from circulation in this way. The LDL binds to specific LDL receptors, which cluster in domains of the plasma membrane known as coated pits (discussed in sub- sequent paragraphs). These domains eventually invaginate to form coated vesicles (Figure 24.39), which pinch off from the plasma membrane and form endosomes (literally “bodies inside” the cell). In the low pH environment of the endosome, the LDL particles dissociate from their receptors. The endosomes then fuse with lyso- somes, and the LDLs are degraded by lysosomal acid lipases. HDLs have much longer life spans in the body (5 to 6 days) than other lipopro- teins. Newly formed HDL contains virtually no cholesterol ester. However, over time, cholesterol esters are accumulated through the action of lecithinϺcholesterol acyltransferase (LCAT), a 59-kD glycoprotein associated with HDLs. Another asso- ciated protein, cholesterol ester transfer protein, transfers some of these esters to VLDL and LDL. Alternatively, HDLs function to return cholesterol and cholesterol esters to the liver. This latter process apparently explains the correlation between high HDL levels and reduced risk of cardiovascular disease. (High LDL levels, on Concentration in Plasma Apoprotein M r (mg/100 mL) Distribution A-1 28,300 90–120 Principal protein in HDL A-2 8,700 30–50 Occurs as dimer mainly in HDL B-48 240,000 Ͻ5 Found only in chylomicrons B100 500,000 80–100 Principal protein in LDL C-1 7,000 4–7 Found in chylomicrons, VLDL, HDL C-2 8,800 3–8 Found in chylomicrons, VLDL, HDL C-3 8,800 8–15 Found in chylomicrons, VLDL, IDL, HDL D 32,500 8–10 Found in HDL E 34,100 3–6 Found in chylomicrons, VLDL, IDL, HDL Adapted from Brown, M., and Goldstein, J., 1987. In Braunwald, E., et al., eds., Harrison’s Principles of Internal Medicine, 11th ed. New York: McGraw-Hill; and Vance, D., and Vance, J.,eds., 1985. Biochemistry of Lipids and Membranes, Menlo Park, CA: Benjamin/Cummings. TABLE 24.2 Apoproteins of Human Lipoproteins (a) (b) FIGURE 24.37 A model for the structure of a typical lipoprotein. (a) A core of cholesterol and cholesteryl esters is surrounded by a phospholipid (monolayer) membrane. Apolipoprotein A-I is modeled here as a long amphipathic ␣-helix, with the nonpolar face of the helix embedded in the hydrophobic core of the lipid particle and the polar face of the helix exposed to solvent. (b) A ribbon diagram of apolipoprotein A-I. (Adapted from Borhani, D.W., Rogers, D.P., Engler, J. A., and Brouillette, C. G., 1997. Crystal structure of truncated human apolipoprotein A-I sug- gests a lipid-bound conformation. Proceedings of the National Academy of Sciences 94:12291–12296.) 24.5 How Are Lipids Transported Throughout the Body? 759 Endoplasmic reticulum Assembly of components into prelipoprotein particles in the ER, then transfer to Golgi Secretory vesicle Golgi Liver cell Extracellular space Golgi processes the particles with additional phospholipids and perhaps also cholesterol and cholesterol esters added Synthesis of apoproteins, phosphatidylcholine, triacylglycerol, cholesterol, cholesterol esters occurs in the endoplasmic reticulum 1 2 3 VLDL Formation of secretory vesicle containing lipoprotein particles 4 The VLDL is released into the circulation 5 FIGURE 24.38 Lipoprotein components are synthesized predominantly in the ER of liver cells. Following assem- bly of lipoprotein particles (red dots) in the ER and processing in the Golgi, lipoproteins are packaged in secre- tory vesicles for export from the cell (via exocytosis) and released into the circulatory system. + LDL LDL LDL receptor Vesicle loses its coating and forms endosome Recycling vesicle LDL receptors bud off and form a small recycling vesicle Endosome formation may or may not include fusion with another vesicle Synthesis of LDL receptors Synthesis of cholesterol Oversupply of cholesterol ACAT Remaining vesicle fuses with lysosome Apoprotein B is degraded by lysosomal protease and released as amino acids Apoprotein B Amino acids Free cholesterol HMG-CoA reductase Inhibits Activates Free cholesterol released Lysosome Cholesterol esters in core are hybridized by ACAT and stored in cell FIGURE 24.39 Endocytosis and degradation of lipoprotein particles.(ACAT is acyl-CoA cholesterol acyltransferase.) 760 Chapter 24 Lipid Biosynthesis the other hand, are correlated with an increased risk of coronary artery and cardio- vascular disease.) The Structure of the LDL Receptor Involves Five Domains The LDL receptor in plasma membranes (Figure 24.40) consists of 839 amino acid residues and is composed of five domains, two of which contain multiple subdo- mains. The N-terminal LDL-binding domain (292 residues) contains seven cysteine- rich repeats, denoted R1 to R7. The next segment (417 residues) contains three epidermal growth factor repeats, as well as a -propellor module. This is followed in the sequence by a 58-residue segment of O-linked oligosaccharides, a 22-residue membrane-spanning segment, and a 50-residue segment extending into the cytosol. The clustering of receptors prior to the formation of coated vesicles requires the presence of this cytosolic segment. Note that the LDL particle binds specifically to the receptor at the fourth and fifth cysteine-rich repeats (R4 and R5). The LDL Receptor -Propellor Displaces LDL Particles in Endosomes Figure 24.39 shows the release of LDL particles in endosomes that pinch off from the plasma membrane when cells take up LDLs. What molecular events trigger the release of LDL particles? A collaboration by three Nobel laureates has provided an answer. Johann Deisenhofer, Michael Brown, and Joseph Goldstein have deter- mined the structure of the extracellular domain of the LDL receptor at pH 5.3, the typical pH inside endosomes. At this low pH, the receptor polypeptide is folded back on itself, with the -propellor domain associated with R4 and R5, the two re- peats that normally bind the LDL particle (Figure 24.41). The implication is that the -propellor displaces the LDL particle in the lower pH environment of the en- dosome. What residues at the interface between the propellor and the R4 and R5 repeats act as the pH sensors? Three histidines at the propellor–R4/R5 interface— His 190 , His 562 , and His 586 —are the likely pH-sensing residues. His 190 lies at the tip of a loop on R5, whereas His 562 and His 586 are on the surface of the propellor domain (Figure 24.41). These three His residues form a cluster at the three-way junction be- tween R4, R5, and the -propellor. Defects in Lipoprotein Metabolism Can Lead to Elevated Serum Cholesterol The mechanism of LDL metabolism and the various defects that can occur therein have been studied extensively by Michael Brown and Joseph Goldstein, who received the Nobel Prize in Physiology or Medicine in 1985. Familial hypercholesterolemia is the term given to a variety of inherited metabolic defects that lead to greatly elevated levels of serum cholesterol, much of it in the form of LDL particles. The general Cys-rich repeats -propellor EGF repeat EGF repeats O-linked oligosaccharide domain 58 residues Transmembrane domain 22 residues Cytosolic domain 50 residues C R7 R2 R1 R3 R4 R5 R6 LDL N FIGURE 24.40 The structure of the LDL receptor.The amino-terminal binding domain is responsible for recognition and binding of LDL apoprotein.The B-100 apolipoprotein of the LDL particle is presumed to bind to the fourth and fifth cysteine-rich repeats (R4 and R5). The O-linked oligosaccharide-rich domain may act as a molecular spacer, raising the binding domain above the glycocalyx.The cytosolic domain is required for aggre- gation of LDL receptors during endocytosis. (a) R4 R5 H 190 (b) (c) R4 R3 R7 R6 R2 R5 -propellor FIGURE 24.41 (a) The cysteine-rich repeats R4 and R5 are the site of LDL particle binding.Each repeat con- tains two loops connected by three disulfide bonds.The second loop in each repeat carries acidic residues (Asp and Glu) and forms a Ca 2ϩ -binding site. (b) The -propellor domain of the LDL receptor. (c) The struc- ture of the LDL receptor extracellular domain at pH 5.3 (similar to that found in endosomes).The -propellor domain is associated with cysteine-rich domains R4 and R5. A cluster of histidines (His 190 , His 562 , and His 586 ) and a variety of hydrophobic and charged interactions mediate the interaction (pdb id ϭ 1N7D). 24.6 How Are Bile Acids Biosynthesized? 761 genetic defect responsible for familial hypercholesterolemia is the absence or dys- function of LDL receptors in the body. Only about half the normal level of LDL re- ceptors is found in heterozygous individuals (persons carrying one normal gene and one defective gene). Homozygotes (with two copies of the defective gene) have few, if any, functional LDL receptors. In such cases, LDLs (and cholesterol) cannot be absorbed, and plasma levels of LDL (and cholesterol) are very high. Typical hetero- zygotes display serum cholesterol levels of 300 to 400 mg/dL, but homozygotes carry serum cholesterol levels of 600 to 800 mg/dL or even higher. There are two possible causes of an absence of LDL receptors—either receptor synthesis does not occur at all, or the newly synthesized protein does not successfully reach the plasma mem- brane due to faulty processing in the Golgi or faulty transport to the plasma mem- brane. Even when LDL receptors are made and reach the plasma membrane, they may fail to function for two reasons. They may be unable to form clusters competent in coated pit formation because of folding or sequence anomalies in the carboxy- terminal domain, or they may be unable to bind LDL because of sequence or fold- ing anomalies in the LDL-binding domain. 24.6 How Are Bile Acids Biosynthesized? Bile acids, which exist mainly as bile salts, are polar carboxylic acid derivatives of cholesterol that are important in the digestion of food, especially the solubilization of ingested fats. The Na ϩ and K ϩ salts of glycocholic acid and taurocholic acid are the principal bile salts (Figure 24.42). Glycocholate and taurocholate are conjugates of HO CH 3 H 3 C COO – CH 3 CH 3 OH H OH HO H 3 N CH 2 COO – H 3 N CH 2 SO 3 – CH 2 O CH 3 CH 3 OH H HO H 3 C C O CH 3 CH 3 OH HO H HO H 3 C C N H COO – HO SO 3 – N H HO CH 3 H 3 C + + OH 7␣-Hydroxycholesterol 7␣-Hydroxylase Cholic acid Glycine Many steps Taurine Glycocholic acid Taurocholic acid Cholesterol FIGURE 24.42 Cholic acid, a bile salt, is synthesized from cholesterol via 7␣-hydroxycholesterol. Conjugation with taurine or glycine produces taurocholic acid and glycocholic acid, respectively. Taurocholate and glyco- cholate are freely water soluble and are highly effective detergents. 762 Chapter 24 Lipid Biosynthesis cholic acid with glycine and taurine, respectively. Because they contain both nonpo- lar and polar domains, these bile salt conjugates are highly effective as detergents. These substances are made in the liver, stored in the gallbladder, and secreted as needed into the intestines. The formation of bile salts represents the major pathway for cholesterol degra- dation. The first step involves hydroxylation at C-7 (Figure 24.42). 7␣-Hydroxylase, which catalyzes the reaction, is a mixed-function oxidase involving cytochrome P-450. Mixed-function oxidases use O 2 as substrate. One oxygen atom goes to hydroxylate the substrate while the other is reduced to water (Figure 24.43). The function of cytochrome P-450 is to activate O 2 for the hydroxylation reaction. Such hydroxy- lations are quite common in the synthetic routes for cholesterol, bile acids, and steroid hormones and also in detoxification pathways for aromatic compounds. Several of these are considered in the next section. 7␣-Hydroxycholesterol is the precursor for cholic acid. 24.7 How Are Steroid Hormones Synthesized and Utilized? Steroid hormones are crucial signal molecules in mammals. (The details of their physiological effects are described in Chapter 32.) Their biosynthesis begins with the desmolase reaction, which converts cholesterol to pregnenolone (Figure 24.44). Desmolase is found in the mitochondria of tissues that synthesize steroids (mainly the adrenal glands and gonads). Desmolase activity includes two hydroxy- lases and utilizes cytochrome P-450. HUMAN BIOCHEMISTRY Steroid 5␣-Reductase—A Factor in Male Baldness, Prostatic Hyperplasia, and Prostate Cancer An enzyme that metabolizes testosterone may be involved in the benign conditions of male-pattern baldness (also known as andro- genic alopecia) and benign prostatic hyperplasia (prostate gland en- largement), as well as potentially fatal prostate cancers. Steroid 5␣-reductases are membrane-bound enzymes that catalyze the NADPH-dependent reduction of testosterone to dihydrotestos- terone (DHT) (see accompanying figure). Two isoforms of 5␣- reductase have been identified. In humans; the type I enzyme pre- dominates in the sebaceous glands of skin and liver, whereas type II is most abundant in the prostate, seminal vesicles, liver, and epi- didymis. DHT is a contributory factor in male baldness and pro- static hyperplasia, and it has also been shown to act as a mitogen (a stimulator of cell division). For these reasons, 5␣-reductase in- hibitors are potential candidates for treatment of these human conditions. Finasteride (see figure) is a specific inhibitor of type II 5␣- reductase. It has been used clinically for treatment of benign pro- static hyperplasia, and it is also marketed under the trade name Propecia by Merck as a treatment for male baldness. Type II 5␣- reductase inhibitors may also be potential therapeutic agents for treatment of prostate cancer. Somatic mutations occur in the gene for type II 5␣-reductase during prostate cancer progression. Because type I 5␣-reductase is the predominant form of the en- zyme in human scalp, the mechanism of finasteride’s promotion of hair growth in men with androgenic alopecia has been uncertain. However, scientists at Merck have shown that whereas type I 5␣-reductase predominates in sebaceous ducts of the skin, type II 5␣-reductase is the only form of the enzyme present in hair follicles. Thus, finasteride’s therapeutic effects may arise from in- hibition of the type II enzyme in the hair follicle itself. OH O CH 3 H CH 3 C NHC(CH 3 ) 3 O CH 3 H N H H H CH 3 O Dihydrotestosterone Finasteride 7␣-Hydroxycholesterol Cholesterol +2 H + H 2 OO 2 Cytochrome P-450 reductase (Flavin-H 2 ) Cytochrome P-450 reductase (Flavin) 7␣-Hydroxylase (Cytochrome P-450) Fe 2+ 7␣-Hydroxylase (Cytochrome P-450) Fe 3+ NADPH + H + NADP + FIGURE 24.43 The mixed-function oxidase activity of 7␣-hydroxylase.