Cholesterol Biosynthesis: Lanosterol to Cholesterol - Journal of

Research: Science and Education edited by

Concepts in Biochemistry

William M. Scovell Bowling Green State University Bowling Green, OH 43403

Cholesterol Biosynthesis: Lanosterol to Cholesterol John M. Risley Department of Chemistry, The University of North Carolina at Charlotte, Charlotte, NC 28223-0001; [email protected]

Cholesterol is an important molecule. It is important biochemically because it maintains membrane fluidity, microdomain structure, and permeability, and it is a precursor to steroid hormones, bile acids, vitamin D, and lipoproteins. In addition to these well-recognized functions, cholesterol has been shown recently to have an essential role in mammalian embryonic development (1). It is important medically because it is correlated with a number of diseases, as, for example, atherosclerosis (2), cholestasis (broad and variable impairment of bile secretion) (3), hypercholesterolemia (4 ), cholesterol gallstone disease (5), and Nieman–Pick type C disease (6 ), on which major research efforts and monies are expended annually. It is important commercially for the pharmaceutical industry in terms of drug development, and for the food industry in terms of “healthy foods” that are advertised either to be “cholesterol-free” or to reduce cholesterol. For example, Zocor (simvastatin) has been advertised to the general public by Merck & Co. as “an effective medicine that along with diet and exercise can significantly lower total cholesterol” (7). And, above the nutrition facts on a box of Kellogg’s Corn Flakes the cereal is touted as a “cholesterol free food”, and on the front of a box of General Mills Cheerios it reads “As part of a heart-healthy diet, the soluble fiber in Cheerios May Reduce Your Cholesterol!” Each cereal “Meets American Heart Association food criteria for saturated fat and cholesterol for healthy people over age 2”. While exogenous cholesterol is considered a major cause for many of the health problems, cholesterol biosynthesis in vivo (endogenous cholesterol) is a target in reducing overall cholesterol in humans (as, for example, by lovastatin, which inhibits HMG-CoA reductase) (2). Considering the significance and interest in cholesterol, it is interesting that discussions of cholesterol biosynthesis in general biochemistry textbooks are restricted to the synthesis of lanosterol from acetate, which is usually discussed in detail. The conversion of lanosterol to cholesterol is most often simply indicated, without elaboration, as a multistep process. The only textbook I know of that presents a pathway of lanosterol to cholesterol is that by Voet and Voet (8), after Rilling and Chayet in 1985 (9). The Biochemical Pathways chart distributed by Roche Molecular Biochemicals (10) gives an abbreviated pathway showing major intermediates, but in general without elaboration; the chart is used by ExPASy with links to a few of the enzymes (11). Map 100 in the Kyoto Encyclopedia of Genes and Genomes (KEGG ) (12) outlines the pathway, naming major intermediates with links to a few of the enzymes, but without elaboration. This situation is unsatisfactory for numerous students, especially those more medically oriented. The research literature on cholesterol biosynthesis is voluminous. A search of “cholesterol biosynthesis” on PubMed

at the National Library of Medicine (13) yielded more than 10,000 citations, while a search of “cholesterol biosynthesis, review” resulted in more than 1000 citations. A significant proportion of the latter are reviews of drug effects on cholesterol biosynthesis. However, I am not aware of a recent review of the pathway from lanosterol to cholesterol that describes in detail the reactions along the line of ref 9. I did find an excellent review published in 1997 that summarizes the cloning of cDNAs or genes encoding the enzymes of sterol biosynthesis for fungi, mammals, and higher plants (14), including brief descriptions of the enzyme reactions for each of the pathways to ergosterol, cholesterol, stigmasterol, and campesterol. Advances have been made since the publication of the review article. For example, the genes for C3 sterol dehydrogenase (C4 decarboxylase) (15) and 3-keto reductase (16 ) have been cloned, completing the elucidation of the pathway of ergosterol biosynthesis in the yeast Saccharomyces cerevisiae. With no recent review from which to teach the conversion of lanosterol to cholesterol, I have used the outline presented in 1985 (9), with supplements obtained in my readings from the research literature describing subsequent information on the enzymes in this pathway. This paper presents the material that I use in teaching this topic. It is divided into five sections: (i) general overview of the pathway, (ii) step-by-step synthesis, (iii) cholesterol biosynthesis in perspective, (iv) cholesterol homeostasis, and (v) inherited diseases of cholesterol biosynthesis. I do not present a comprehensive review of the literature in this area, but I have included recent relevant references from which a more comprehensive study may be pursued if desired. Overview of the Pathway The anabolic pathway for the conversion of lanosterol to cholesterol is shown in Figure 1. The numbering scheme for lanosterol is shown in Figure 2. The biosynthesis of cholesterol from lanosterol is a 19step process. It requires nine different enzymes; two enzymes catalyze multiple steps and three are utilized two times in the pathway. Three methyl groups in lanosterol (Fig. 2: C30, C31, C32) are oxidized and removed as formic acid and two carbon dioxide molecules. Two mechanisms are utilized in these reactions: a cytochrome P450 and formation of a βketoacid. The other reactions in the pathway are five reduction reactions using NAD(P)H, one dehydrogenation (desaturation) reaction using NADH/O2, and one isomerization. Two mechanisms are utilized in the reduction reactions: two ketoreductions and three olefinic reductions involving carbocation intermediates. A radical mechanism is utilized in the dehydrogenation (desaturation) reaction, and formation of a carbocation intermediate is utilized in the isomerization reaction.

JChemEd.chem.wisc.edu • Vol. 79 No. 3 March 2002 • Journal of Chemical Education

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Research: Science and Education

Figure 1. The pathway for the 19-step conversion of lanosterol to cholesterol.

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Journal of Chemical Education • Vol. 79 No. 3 March 2002 • JChemEd.chem.wisc.edu


Step-by-Step Synthesis

22

Reduction of 5 to 6 The reduction of cholestatriene (5) to 14-demethyllanosterol (4,4′-dimethylzymosterol; 4,4′-dimethylcholesta8,24-dien-3β-ol) (6) is catalyzed by ∆14-sterol reductase (sterol ∆14-reductase; steroid-14-reductase; ∆8,14-sterol ∆14-reductase) (26, 27 ). The enzyme requires NADPH and the mechanism involves formation of a carbocation intermediate (Fig. 4). A proton from the aqueous solution (or from a general acid in the enzyme active site) protonates at C15 to form a carbocation, which is stabilized by the active site of the enzyme and by the adjacent double bond. The enzyme directs transfer of a hydride from NADPH to C14 that completes the reduction reaction.

20

18

In the following discussion of the 19-step reaction, I give the various names used in the literature for the enzyme, substrates, and products in each step.

Oxidative Demethylation of 1 to 5 The oxidative demethylation of lanosterol (4,4′,14α-trimethyl-5α-cholesta-8(9),24-dien-3β-ol) (1) to cholestatriene (4,4′-dimethyl-5α -cholesta-8(9),14,24-trien-3β -ol; 4,4′dimethylcholesta-8,14,24-trien-3β-ol) (5) is catalyzed by lanosterol 14α-methyl demethylase cytochrome P450 (lanosterol 14α-methyl demethylase; 14α-methyl demethylase; lanosterol 14α-demethylase) (17–22). The 14α-methyl group, which protrudes into the α face of the sterol ring system, is the first group to be removed; its removal is essential for membrane and regulatory functions of the resulting sterols (14). The enzyme, a cytochrome P450, requires the C3-hydroxy group, 14αmethyl group, and ∆8(9) bond for binding and catalysis (22); it also requires three NADPH coenzymes and three O2 molecules. Upon substrate binding the heme shifts from a low-spin to a high-spin state (22). The enzyme catalyzes three successive monooxygenation reactions: (i) lanosterol (1) to 3β-hydroxylanost-8-en-32-ol (lanost-8-en-3β,32-diol) (2), (ii) 3β-hydroxylanost-8-en-32-ol (2) to 3β-hydroxylanost-8-en-32-al (3), and (iii) 3β-hydroxylanost-8-en-32-al (3) to 14α-formyloxy-lanost-8-en-3β-ol (4) to cholestatriene (5). The third oxidation has been proposed to proceed by peroxy attack, a Baeyer–Villiger rearrangement, and deformylation by a base rearrangement (17), or by peroxy attack, homolytic cleavage, deformylation by fragmentation, and disproportionation (20). (Deformylation by fragmentation has been termed a radicalar mechanism [14].) The human cDNA clone has been reported (23) and a model of the Candida albicans enzyme has been proposed (24). The crystal structure of the enzyme from Mycobacterium tuberculosis was recently published (PDB ID 1E9X and 1EA1) (25). This is the first enzyme in this pathway for which there is a crystal structure. The enzyme is inhibited by compounds, such as miconazole and ketoconazole (lipophilic imidazoles) (Fig. 3), that coordinate through the imidazole nitrogen to the heme iron of the cytochrome P450 and block binding of O2 (14, 25). The result is an accumulation of dihydrolanosterol (the C24 reduction product of lanosterol).

24

27

21

11 9

19 2 β 3

HO

β

1 4

31

12

13 14

8

6

26

16 15

α

10 5

17

25

23

32

7

α 30

Figure 2. The numbering scheme for lanosterol.

Cl N N

CH2

CH

O

CH2

Cl

Cl miconazole Cl

O

N

N

COCH3

O

N N

CH2

C

O Cl

ketoconazole

Cl

Figure 3. The structures for two inhibitors of lanosterol 14α-methyl demethylase cytochrome P450.

5

HO

H* HO

Hw 6

Hw+ NADPH*

+

Hw

HO

Figure 4. Reduction of the C14 double bond by ∆14-sterol reductase.


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Oxidation of 6 to 9 The oxidation of 4,4′-dimethylzymosterol (6) to 4methylzymosterol-4-carboxylic acid (4β-methyl-4α-carboxycholesta-8,24-dien-3β-ol) (9) is catalyzed by sterol-4α-methyloxidase (C4 sterol methyl oxidase; C4 methyl sterol oxidase; 4-methyl sterol oxidase; C4 methyl oxidase; C4-methyloxidase) (28, 29). The enzyme is specific for oxidation of the C4α methyl group. It requires three NADH coenzymes and three O2 molecules. The reducing equivalents from NADH are acquired by NADH:cytochrome b5 reductase and cytochrome b5. The enzyme catalyzes three successive (monooxygenation) oxidation reactions: (i) 4,4′-dimethylzymosterol (6) to 4-methyl4-hydroxymethylzymosterol (4β-methyl-4α-hydroxymethylcholesta-8,24-dien-3β-ol) (7), (ii) 4-methyl-4-hydroxymethylzymosterol (7) to 4-methylzymosterol-4-carboxaldehyde (4βmethyl-4α-formyl-cholesta-8,24-dien-3β-ol) (8), and (iii) 4-methylzymosterol-4-carboxaldehyde (8) to 4-methylzymosterol-4-carboxylic acid (9). The human cDNA clone has been mapped to chromosome 4q32-34, and histidine-rich clusters are suggested to be involved in the binding of oxodiiron (Fe–O–Fe) (29).

NAD+ H O H

Oxidation of 11 to 14 The oxidation of 4 α -methylzymosterol (11) to zymosterol-4α-carboxylic acid (4α-carboxy-cholesta-8,24dien-3β-ol) (14) is catalyzed by sterol-4α-methyl-oxidase, the same enzyme that catalyzes the oxidation of 6 to 9. It catalyzes three successive (monooxygenation) oxidation reactions: (i) 4 α -methylzymosterol (11) to 4 α -hydroxymethylzymosterol (4α -hydroxymethyl-cholesta-8,24-dien-3β-ol) (12), (ii) 4α-hydroxymethylzymosterol (12) to zymosterol4α-carboxaldehyde (4α-formyl-cholesta-8,24-dien-3β-ol) (13), and (iii) zymosterol-4 α -carboxaldehyde (13) to zymosterol-4α-carboxylic acid (14). Oxidative Decarboxylation of 14 to 15 The oxidative decarboxylation of zymosterol-4α-carboxylic acid (14) to zymosterone (15) is catalyzed by C3 sterol dehydrogenase (C4 decarboxylase), which is the same enzyme 380

O HO

CO2

O

C O

O

H O

10 H

Figure 5. Oxidative decarboxylation and enolization epimerization of the 3β-hydroxycarboxylic acid by C3 sterol dehydrogenase (C4 decarboxylase).

Oxidative Decarboxylation and Enolization Epimerization of 9 to 10 The oxidative decarboxylation and enolization epimerization of 4-methylzymosterol-4-carboxylic acid (9) to 4-methylzymosterone (3-keto-4-methylzymosterol) (10) is catalyzed by C3 sterol dehydrogenase (C4 decarboxylase) (4α-oic acid decarboxylase; C4 decarboxylase) (15). The enzyme requires an NAD+ coenzyme. The enzyme catalyzes oxidation of 9 (Fig. 5) by transfer of the 3α hydrogen as a hydride to NAD+, leading to a 3-ketocarboxylic acid intermediate that undergoes decarboxylation to give an enol. The enzyme directs the epimerization from the enol that moves the former 4β-methyl group to the 4α-methyl position. Reduction of 10 to 11 The reduction of 4-methylzymosterone (10) to 4αmethylzymosterol (4α-methyl-cholesta-8,24-dien-3β-ol) (11) is catalyzed by 3-keto reductase (3-ketosterol reductase; steroid3-ketoreductase) (16 ). The enzyme, which requires an NAD(P)H coenzyme, catalyzes the 3-ketoreduction to convert the 3-keto group to the β-hydroxy sterol.

9

C O –

HO

H

16

H* Hw+

Hw +

HO

H H*

H*+ HO

17

Figure 6. The isomerization reaction catalyzed by ∆ 8-∆7 sterol isomerase.

that catalyzes the oxidative decarboxylation and enolization epimerization of 9 to 10. The enzyme catalyzes oxidation of 14 to a 3-ketocarboxylic acid intermediate that undergoes decarboxylation to give an enol, which undergoes tautomerization.

Reduction of 15 to 16 The reduction of zymosterone (15) to zymosterol (5αcholesta-8,24-dien-3 β -ol; 5 α -cholest-8-en-3 β -ol; ∆ 8,24cholestadien-3β-ol; ∆8-cholestenol) (16) is catalyzed by 3-keto reductase, the same enzyme that catalyzes the reduction of 10 to 11. The enzyme catalyzes the 3-ketoreduction to convert the 3-keto group to the β-hydroxy sterol.



HO Lanthosterol HO

NADH O2

Desmosterol NADPH NAD(P)H

HO 7-Dehydrodesmosterol

Isomerization of 16 to 17 The isomerization of zymosterol (16) to lanthosterol (5αcholesta-7,24-dien-3 β -ol; 5 α -cholest-7-en-3 β -ol; ∆ 7,24cholestadien-3β-ol; cholest-7-en-3β-ol) (17) is catalyzed by ∆8-∆7 sterol isomerase (sterol ∆8-isomerase; 3β-hydroxysteroid-∆8∆7-isomerase; cholestenol ∆-isomerase; ∆7-cholestenol ∆7-∆8isomerase, EC 5.3.3.5) (30–33). The enzyme is also an emopamil-binding protein and a sigma factor. The human cDNA clone has been reported (30). The enzyme catalyzes isomerization of the double bond (Fig. 6) by protonation at C9 by a proton from the aqueous solution (or from a general acid in the enzyme active site) to form a carbocation intermediate, which is stabilized by the active site of the enzyme. The enzyme directs subsequent abstraction of the 7β H to give the product. This isomerization is the only reversible reaction in the 19 steps of biosynthesis of cholesterol from lanosterol (33).

HO Cholesterol

Figure 7. The position in the pathway of C24 reduction has most often been placed as the last step in the conversion of lanosterol to cholesterol. In this position, the pathway from lanthosterol to cholesterol is shown.

H*

18

HO H H

H

HO

H

H

H*

19

HO

Reduction of 17 to 18 The reduction of lanthosterol (17) to lathosterol (5αcholest-7-en-3β-ol; ∆7-cholesten-3β-ol) (18) is catalyzed by sterol ∆24-reductase (24-reductase; 3β-hydroxysterol ∆24-reductase) (34). The exact location in the pathway for the C24 reduction is not known because the enzyme catalyzes the reduction of the double bond at each step in the pathway (e.g., see conversion of 1 to 5). The position of the C24 reduction has most often been placed as the last step in the pathway so that lanthosterol is converted first to 7-dehydrodesmosterol and then to desmosterol before final reduction of C24 to cholesterol (Fig. 7). Bae and Paik (34) reported that the relative rates of reduction by sterol ∆24-reductase were 1.0 for lanthosterol (17), 0.34 for zymosterol (16), 0.31 for desmosterol, and 0.06 for lanosterol (1); lanthosterol is more easily reduced than desmosterol or zymosterol. On the basis of their proposal from this study, I have placed the C24 reduction at this location in the pathway. The enzyme requires an NAD(P)H coenzyme and presumably catalyzes the reduction by a carbocation intermediate.

*

H

Figure 8. Lathosterol oxidase catalyzes the formation of the C5 double bond.

Hw

19

HO

20 +

Hw

HO

H*

NADPH*

Hw

HO

+

Figure 9. The last step in the pathway is reduction of the C7 double bond by 7-dehydrocholesterol reductase.

Dehydrogenation (Desaturation) of 18 to 19 The dehydrogenation (desaturation) of lathosterol (18) to 7-dehydrocholesterol (cholesta-5,7-dien-3β-ol) (19) is catalyzed by lathosterol oxidase (lathosterol 5-desaturase; ∆7-sterol ∆5reductase; ∆7-sterol 5-reductase; ∆7-sterol-C5(6)-desaturase; sterol 5α,6α-desaturase; sterol ∆5 desaturase; ∆5-dehydrogenase; C5-sterol desaturase; 5α-cholest-7-en-3β-ol:O2 ∆5-oxidoreductase, EC 1.3.3.2) (35, 36 ). The enzyme requires an NAD(P)H coenzyme (NADH is better than NADPH) and an O2 molecule; the reducing equivalents from NADH are acquired by NADH:cytochrome b5 reductase and cytochrome b5. The enzyme contains histidine-rich motifs that would provide the ligands for a presumed catalytic Fe center (35). It catalyzes the stereospecific dehydrogenation (desaturation) of the 5α and 6α hydrogen atoms in a cis abstraction. A study indicated an asynchronous scission of the two C–H bonds (Fig. 8) where the C6α H is removed first to form a radical intermediate, and then the C5α H is removed, rather than a concerted desaturation mechanism (36 ). The human cDNA clone has been reported (37 ).


381


Reduction of 19 to 20 The reduction of 7-dehydrocholesterol (19) to cholesterol (20) is catalyzed by 7-dehydrocholesterol reductase (∆7-sterol reductase; 3β-hydroxysterol ∆7-reductase; 7-dehydrocholesterol ∆7-reductase; cholesterol:NADP + ∆7-oxidoreductase, EC 1.3.1.21) (38, 39). The enzyme requires an NADPH coenzyme. It catalyzes ∆7-reduction (Fig. 9) by protonation at C8β by a proton from aqueous solution (or from a general acid in the enzyme active site) to form a carbocation intermediate that is stabilized by the active site of the enzyme and by the adjacent double bond. The enzyme directs hydride transfer from NADPH to C7α that completes the reduction reaction. The human cDNA clone has been reported (38) and mapped to chromosome 11q12-13 (40). The 19-step conversion of lanosterol to cholesterol requires nine different enzymes. It has been proposed to occur in both the endoplasmic reticulum and the peroxisome compartments of the cell (41). trans-1,4-Bis-(2-chlorobenzylaminomethyl)cyclohexane dihydrochloride (AY-9944) is an inhibitor of ∆14-sterol reductase (conversion of 5 to 6), ∆8–∆7 sterol isomerase (conversion of 16 to 17), and 7-dehydrocholesterol reductase (conversion of 19 to 20) with application in hyperlipidemia. trans-2-[4-(1,2-Diphenylbuten-1-yl)phenoxy]N,N-dimethylethylamine (tamoxifin), a chemotherapeutic agent used in the treatment of breast cancer, is an inhibitor of ∆8–∆7 sterol isomerase. Cholesterol Biosynthesis in Perspective The biosynthesis of cholesterol from lanosterol is not unlike other metabolic pathways in that the enzymes catalyze a series of oxidation and reduction reactions along with carbon– carbon bond cleavage, epimerization, isomerization, and dehydrogenation. The five reduction reactions (5 → 6, 10 → 11, 15 → 16, 17 → 18, 19 → 20) utilize reduced flavoenzymes (NAD[P]H) that are common in metabolic pathways (42). The epimerization (part of 9 → 10) and isomerization (16 → 17) reactions are also observed in other metabolic pathways (42). Removal of the three methyl groups from lanosterol (C30, C31, C32) requires carbon–carbon bond cleavage. To accomplish this, the methyl groups are oxidized to carboxylic acids and eliminated as a formic acid (4 → 5) and two carbon dioxides (part of 9 → 10, 14 → 15). Three enzymes catalyze the demethylation reactions. A cytochrome P450 enzyme is used for one complete oxidative decarboxylation of C32 (1 → 5), and two enzymes are used in the oxidation (6 → 9, 11 → 14) and oxidative decarboxylation (9 → 10, 14 → 15) reactions of C30 and C31. Oxidations catalyzed by cytochrome P450 enzymes are common (43–45), but oxidations ending in carbon–carbon bond cleavage appear unique to certain steroid-metabolizing cytochrome P450 enzymes (43, 44), including lanosterol 14α-methyl demethylase cytochrome P450 here. Thus, this type of carbon–carbon bond cleavage is not widely seen in metabolic pathways. On the other hand, the oxidation reactions of C30 and C31 are thought to involve non-heme oxo-diiron (29). The mechanism for oxidation by these binuclear nonheme iron centers is believed to be formally equivalent to that of cytochrome P450 (36, 42, 46 ), and enzymes utilizing non-heme oxo-diiron centers are being more widely identified (36, 46 ). But the reaction here does not end in carbon–carbon 382

bond cleavage. Instead, a second enzyme catalyzes oxidation of the β-hydroxyacid to a β-ketoacid, which undergoes decarboxylation. Decarboxylation via a β-ketoacid is more familiar in metabolic pathways, as, for example, in the citric acid cycle where isocitrate dehydrogenase catalyzes oxidative decarboxylation of isocitrate (via oxalosuccinate) to α-ketoglutarate, and in the oxidative phase of the pentose phosphate pathway where 6-phosphogluconate dehydrogenase catalyzes oxidative decarboxylation of 6-phosphogluconate (via 6-phospho-3-oxogluconate) to ribulose-5-phosphate. Therefore, the biosynthetic pathway for cholesterol from lanosterol displays two significantly different mechanisms for carbon–carbon bond cleavage; the mechanisms are related via the oxidation reactions using iron centers. Finally, the dehydrogenation (desaturation) reaction (18 → 19) is not catalyzed by a flavoenzyme, although these are widespread in metabolic pathways (e.g., succinate dehydrogenase in the citric acid cycle, fatty acyl-CoA dehydrogenase in the β-oxidation of fatty acids, and dihydroorotate oxidase in pyrimidine nucleotide biosynthesis). Instead, a reduced flavin (NAD[P]H) and O2 are involved, presumably with a nonheme iron (oxo-diiron) center. These non-heme iron enzymes participate in the biosynthesis of unsaturated fatty acids as fatty acyl-CoA desaturases (e.g., ∆6-desaturase, ∆9-stearoylCoA desaturase, ∆11-myristoyl-CoA desaturase, and ∆12-oleylCoA desaturase) (36 ). Thus, this type of dehydrogenation may be primarily associated with lipid metabolism. Cholesterol Homeostasis Much has been learned about cholesterol homeostasis, although a complete picture has not yet emerged. Central to cholesterol homeostasis are the sterol regulatory elementbinding proteins (SREBPs) (particularly SREBP-2, but also SREBP-1c), which are under sterol regulation. SREBP-1a and -1c primarily regulate fatty acid biosynthesis (47–49). When intracellular cholesterol levels decline, SREBP-cleavage activating protein (SCAP) forms a complex with SREBP that mediates the interaction of SREBP with site-1-protease (S1P). A recent study indicated that modifications of the N-linked carbohydrate moieties on SCAP are important for this to occur (50). S1P is a membrane-bound subtilisin-related serine protease (51) that catalyzes the first proteolytic reaction on SREBP. Site-2-protease (S2P), a member of the family of zinc metalloproteases (51), catalyzes hydrolysis of the second proteolytic reaction on SREBP, which releases a soluble transcription factor (the amino-terminal fragment of SREBP). The soluble transcription factor enters the nucleus, where (usually with a second transcription factor such as Sp-1 or NF-1) it binds to direct repeat sterol response elements (SRE) in promotor regions, activating transcription of sterol-regulated cholesterogenic genes. Genes under this regulatory control include, at least, LDL receptor, HMG-CoA synthase, HMG-CoA reductase, farnesyl diphosphate synthase, squalene synthase, lanosterol synthase, and lanosterol 14α-methyl demethylase cytochrome P450. Activation of these genes restores intracellular cholesterol levels. When intracellular cholesterol levels build up, the activation of SREBPs by proteolysis is suppressed, which decreases gene transcription for cholesterol biosynthesis. Cholesterol itself, when added to cultured cells, only weakly suppresses



cholesterol biosynthesis. However, when 25-hydroxycholesterol (cholest-5-ene-3β,25-diol), an oxysterol (52) synthesized by cholesterol 25-hydroxylase (53), is added to cultured cells there is a rapid decrease in the activation of SREBPs and a consequent rapid decrease in cholesterol biosynthesis. If 25hydroxycholesterol is the regulatory agent, the recent study indicates that the sterol blocks modification of the N-linked carbohydrate moieties of SCAP, which precludes an interaction between S1P and SREBP (50). Oxysterols (formed by hydroxylation of the side chain of cholesterol) are modulators of numerous processes (52). They are important in cholesterol homeostasis in peripheral tissues such as brain and kidney where excess cholesterol is converted to oxysterols that are more hydrophilic than cholesterol. These are transported to the liver for conversion to bile acids, which is under control of nuclear orphan receptors (54). Bile acids are synthesized by two biosynthetic pathways: from cholesterol (the “classical” pathway) and from oxysterols (a newly described pathway) (54).

malformation of the face, limb abnormalities, ambiguous genitalia, and mental retardation. There are two phenotypes: type I (mild) and type II (severe). The incidence of this autosomal recessive disorder is estimated to be 1 in 20,000 births. It may be the second most common autosomal recessive disorder in the white population of North Americans (after cystic fibrosis). Approximately three-quarters of an issue of the American Journal of Medical Genetics was recently devoted to SLOS (64 ). Recognition of cholesterol’s essential role in mammalian embryonic development (1) established SLOS as the prototypical developmental disorder (62).

Inherited Diseases of Cholesterol Biosynthesis

1. Farese, R. V. Jr.; Herz, J. Trends Genet. 1998, 14, 115. 2. Atherosclerosis and Coronary Artery Disease; Fuster, V.; Ross, R.; Topol, E. J., Eds.; Lippincott-Raven: Philadelphia, 1996. 3. Chowdhury, J. R.; Walkoff, A. W.; Chowdhury, N. R.; Arias, I. M. In The Metabolic and Molecular Bases of Inherited Disease, 7th ed.; Scriver, C. R.; Beaudet, A. L.; Sly, W. S.; Valle, D., Eds.; McGraw-Hill: New York, 1995; p 2161. 4. Goldstein, J. L.; Hobbs, H. H.; Brown, M. S. In The Metabolic and Molecular Bases of Inherited Disease, 7th ed.; Scriver, C. R.; Beaudet, A. L.; Sly, W. S.; Valle, D., Eds.; McGrawHill: New York, 1995; p 1981. 5. Way, L. W. In Cecil Textbook of Medicine, 16th ed.; Wyngaarden, J. B.; Smith, L. H., Eds.; Saunders: Philadelphia, 1982; p 752. 6. Pentchev, P. G.; Vanier, M. T.; Suzuki, K.; Patterson, M. C. In The Metabolic and Molecular Bases of Inherited Disease, 7th ed.; Scriver, C. R.; Beaudet, A. L.; Sly, W. S.; Valle, D., Eds.; McGraw-Hill: New York, 1995; p 2625. 7. U.S. News and World Report 2001, 130 (10, Mar 12), 14. 8. Voet, D.; Voet, J. G. Biochemistry, 2nd ed.; Wiley: New York, 1995; p 699. 9. Rilling, H. C.; Chayet, L. T. In Sterols and Bile Acids; Danielsson, H.; Sjövall, J., Eds.; Elsevier: New York, 1985; p 33. 10. Biochemical Pathways, 3rd ed.; Michal, G., Ed.; Boehringer Mannheim: Mannheim, Germany, 1993. 11. ExPASy Molecular Biology Server; http://www.expasy.ch (accessed Dec 2001). 12. Bioinformatics Center, Institute for Chemical Research, Kyoto University. KEGG: Kyoto Encyclopedia of Genes and Genomes; http://www.genome.ad.jp/kegg/ (accessed Dec 2001). 13. National Center for Biotechnology Information Home Page; http://www.ncbi.nlm.nih.gov (accessed Dec 2001). 14. Bach, T. J.; Benveniste, P. Prog. Lipid Res. 1997, 36, 197. 15. Gachotte, D.; Barbuch, R.; Gaylor, J.; Nickel, E.; Bard, M. Proc. Natl. Acad. Sci. USA 1998, 95, 13794. 16. Gachotte, D.; Sen, S. E.; Eckstein, J.; Barbuch, R.; Krieger, M.; Ray, B. D.; Bard, M. Proc. Natl. Acad. Sci. USA 1999, 96, 12655. 17. Fischer, R. T.; Trzaskos, J. M.; Magolda, R. L.; Ko, S. S.; Brosz, C. S.; Larsen, B. J. Biol Chem. 1991, 266, 6124.

Deficiencies in four enzymes of cholesterol biosynthesis from lanosterol have been reported to give rise to diseases. Mutations have been identified in the genes for three of these enzymes. Mutations in C3 sterol dehydrogenase (C4 decarboxylase) (conversion of 9 to 10, 14 to 15) give rise to CHILD (congenital hemidysplasia, ichthyosis, and limb defects) syndrome (55) (MIM 308050 [56 ]). CHILD syndrome is characterized by dry, thickened, scaly skin lesions (“fishskin”) on one side of the body; limb defects; calcification points on cartilage structures; and internal organ anomalies. Mutations in ∆8–∆7 sterol isomerase (conversion of 16 to 17) give rise to two forms of chondrodysplasia punctata (CDP) (57–59). CDP describes calcification points on cartilage structures and disorders of bone growth and structure associated with skeletal dwarfism. One form of CDP is Conradi–Hünermann syndrome (CDPX2) (MIM 302960 [56 ]), which arises by mutations in ∆8–∆7 sterol isomerase (58). It resembles CHILD syndrome, but the dry, thickened, scaly skin lesions are on both sides of the body in an asymmetric pattern. Because CHILD syndrome and CDPX2 have similar clinical features, a patient diagnosed with CHILD syndrome was reported to have a mutation in the ∆8–∆7 sterol isomerase gene (59). Thus CHILD syndrome and CDPX2 were proposed to result from allelic mutations in the ∆8–∆7 sterol isomerase gene. However, because CHILD syndrome was shown to arise by mutations in C3 sterol dehydrogenase (C4 decarboxylase) (55), the mutation in ∆8–∆7 sterol isomerase (59) may be another form of CDP. Therefore, CHILD syndrome is distinct from Conradi–Hünermann (with unilateral skin lesions and bilateral skin lesions). A deficiency in sterol ∆24-reductase (conversion of 17 to 18) activity leading to an accumulation of desmosterol has been proposed to give rise to desmosterolosis (60). This disease is characterized by abnormally large cranial capacity, cleft palate, ambiguous genitalia, and short limbs. Mutations of 7-dehydrocholesterol reductase (conversion of 19 to 20) give rise to Smith–Lemli–Opitz syndrome (SLOS) (40, 61–63) (MIM 270400 [56 ]). SLOS is characterized by abnormal smallness of the head, poor growth, cleft palate,

Acknowledgments I wish to thank James C. Crosthwaite, Thomas W. Mattingly Jr., and Joanna K. Krueger for a critical reading of the manuscript. Literature Cited


383

Research: Science and Education 18. Trzaskos, J. M.; Ko, S. S.; Magolda, R. L.; Favata, M. F.; Fischer, R. T.; Stam, S. H.; Johnson, P. R.; Gaylor, J. L. Biochemistry 1995, 34, 9670. 19. Trzaskos, J. M.; Fischer, R. T.; Ko, S. S.; Magolda, R. L.; Stam, S.; Johnson, P.; Gaylor, J. L. Biochemistry 1995, 34, 9677. 20. Shyadehi, A. Z.; Lamb, D. C.; Kelly, S. L.; Kelly, D. E.; Schunck, W.-H.; Wright, J. N.; Corina, D.; Akhtar, M. J. Biol. Chem. 1996, 271, 12445. 21. Lamb, D. C.; Kelly, D. E.; Schunck, W.-H.; Shyadehi, A. Z.; Akhtar, M.; Lowe, D. J.; Baldwin, B. C.; Kelly, S. L. J. Biol. Chem. 1997, 272, 5682. 22. Bellamine, A.; Mangla, A. T.; Nes, W. D.; Waterman, M. R. Proc. Natl. Acad. Sci. USA 1999, 96, 8937. 23. Strömstedt, M.; Rozman, D.; Waterman, M. R. Arch. Biochem. Biophys. 1996, 329, 73. 24. Ji, H.; Zhang, W.; Zhou, Y.; Zhang, M.; Zhu, J.; Song, Y.; Lü, J.; Zhu, J. J. Med. Chem. 2000, 43, 2493. 25. Podust, L. M.; Poulos, T. L.; Waterman, M. R. Proc. Natl. Acad. Sci. USA 2001, 98, 3068. 26. Kim, C.-K.; Jeon, K.-I.; Lim, D.-M.; Johng, T.-N.; Trzasdos, J. M.; Gaylor, J. L.; Paik, Y.-K. Biochim. Biophys. Acta 1995, 1259, 39. 27. Silve, S.; Dupuy, P. H.; Ferrara, P.; Loison, G. Biochim. Biophys. Acta 1998, 1392, 233. 28. Bard, M.; Bruner, D. A.; Pierson, C. A.; Lees, N. D.; Biermann, B.; Frye, L.; Koegel, C.; Barbuch, R. Proc. Natl. Acad. Sci. USA 1996, 93, 186. 29. Li, L.; Kaplan, J. J. Biol. Chem. 1996, 271, 16927. 30. Hanner, M.; Moebius, F. F.; Weber, F.; Grabner, M.; Striessnig, J.; Glossmann, H. J. Biol. Chem. 1995, 270, 7551. 31. Silve, S.; Dupuy, P. H.; Labit-Lebouteiller, C.; Kaghad, M.; Chalon, P.; Rahier, A.; Taton, M.; Lupker, J.; Shire, D.; Loison, G. J. Biol. Chem. 1996, 271, 22434. 32. Moebius, F. F.; Soellner, K. E. M.; Fiechtner, B.; Huck, C. W.; Bonn, G.; Glossmann, H. Biochemistry 1999, 38, 1119. 33. Bae, S.-H.; Seong, J.; Paik, Y.-K. Biochem. J. 2001, 353, 689. 34. Bae, S.-H.; Paik, Y.-K. Biochem. J. 1997, 326, 609. 35. Taton, M.; Husselstein, T.; Benveniste, P.; Rahier, A. Biochemistry 2000, 39, 701. 36. Rahier, A. Biochemistry 2001, 40, 256. 37. Matsushima, M.; Inazawa, J.; Takahashi, E.; Suzumori, K.; Nakamura, Y. Cytogenet. Cell Genet. 1996, 74, 252. 38. Moebius, F. F.; Fitzky, B. U.; Lee, J. N.; Paik, Y.-K.; Glossmann, H. Proc. Natl. Acad. Sci. USA 1998, 95, 1899. 39. Bae, S.-H.; Lee, J. N.; Fitzky, B. U.; Seong, J.; Paik, Y.-K. J. Biol. Chem. 1999, 274, 14624. 40. Wassif, C. A.; Maslen, C.; Kachilele-Linjewile, S.; Lin, D.;

384

41. 42. 43. 44. 45. 46. 47. 48. 49. 50. 51. 52. 53. 54. 55. 56. 57. 58.

59. 60.

61.

62.

63. 64.

Linck, L. M.; Connor, W. E.; Steiner, R. D.; Porter, F. D. Am. J. Hum. Genet. 1998, 63, 55. Krisans, S. K. Ann. N.Y. Acad. Sci. 1996, 804, 142. Kyte, J. Mechanism in Protein Chemistry; Garland: New York, 1995. Cytochrome P450: Structure, Mechanism and Biochemistry, 2nd ed.; Ortiz de Montellano, P. R., Ed.; Plenum: New York, 1995. Sono, M.; Roach, M. P.; Coulter, E. D.; Dawson, J. H. Chem. Rev. 1996, 96, 2841. Newcomb, M.; Toy, P. H. Acc. Chem. Res. 2000, 33, 449. Wallar, B. J.; Lipscomb, J. D. Chem. Rev. 1996, 96, 2625. Brown, M. S.; Goldstein, J. L. Cell 1997, 89, 331. Ridgway, N. D.; Byers, D. M.; Cook, H. W.; Storey, M. K. Prog. Lipid Res. 1999, 38, 337. Osborne, T. F. J. Biol. Chem. 2000, 275, 32379. Nohturfft, A.; DeBose-Boyd, R. A.; Scheek, S.; Goldstein, J. L.; Brown, M. S. Proc. Natl. Acad. Sci. USA 1999, 96, 11235. Brown, M. S.; Goldstein, J. L. Proc. Natl. Acad. Sci. USA 1999, 96, 11041. Schroepfer, G. J. Jr. Physiol. Rev. 2000, 80, 361. Lund, E. G.; Kerr, T. A.; Sakai, J.; Li, W.-P.; Russell, D. W. J. Biol. Chem. 1998, 273, 34316. Russell, D. W. Cell 1999, 97, 539. König, A.; Happle, R.; Bornholdt, D.; Engel, H.; Grzeschik, K.-H. Am. J. Med. Genet. 2000, 90, 339. Online Mendelian Inheritance in Man (OMIM) Home Page; http://www.ncbi.nlm.nih.gov/omim (accessed Dec 2001). Kelley, R. I.; Wilcox, W. G.; Smith, M.; Kratz, L. E.; Moser, A.; Rimoin, D. S. Am. J. Med. Genet. 1999, 83, 213. Braverman, N.; Lin, P.; Moebius, F. F.; Obie, C.; Moser, A.; Glossmann, H.; Wilcox, W. R.; Rimoin, D. L.; Smith, M.; Kratz, L. Nat. Genet. 1999, 22, 291. Grange, D. K.; Kratz, L. E.; Braverman, N. E.; Kelley, R. I. Am. J. Med. Genet. 2000, 90, 328. FitzPatrick, D. R.; Keeling, J. W.; Evans, M. J.; Kan, A. E.; Bell, J. E.; Porteous, M. E. M.; Mills, K.; Winter, R. M.; Clayton, P. T. Am. J. Med. Genet. 1998, 75, 145. Fitzky, B. U.; Witsch-Baumgartner, M.; Erdel, M.; Lee, J. N.; Paik, Y.-K.; Glossmann, H.; Utermann, G.; Moebius, F. F. Proc. Natl. Acad. Sci. USA 1998, 95, 8181. Waterham, H. R.; Wijburg, F. A.; Hennekam, R. C. M.; Vreken, P.; Poll-The, B. T.; Dorland, L.; Duran, M.; Jira, P. E.; Smeitink, J. A. M.; Wevers, R. A.; Wanders, R. J. A. Am. J. Hum. Genet. 1998, 63, 329. Yu, H.; Tint, G. S.; Salen, G.; Patel, S. B. Am. J. Med. Genet. 2000, 90, 347. Am. J. Med. Genet. 1997, 68 (3).


Cholesterol Biosynthesis: Lanosterol to Cholesterol - Journal of

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