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Steroid Synthesis

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Sage Arbor

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Transcript of Steroid Synthesis

17-α-Hydroxylase
(CYP17)
steroid type
Prezi viewable at

Cholesterol
Progesterone
Estradiol
Aldosterone
Cortisol
Testosterone
mineralocorticoid
↑ Na absorption
↑ blood pressure & fluid volume
produced in zona glomerulosa of adrenal cortex
ALDehyde + O + STERONE
ESTRAne + DIOL (2 -oh's)
CHOLE (bile) + STEROs (solid)
dominant glucocorticoid ( GLUCOse + CORTex + sterOID )
synthesized in zona fasciculata of adrenal cortex
↑ blood pressure, ↑Na uptake
17-α-Hydroxyprogesterone
11-Deoxycorticosterone
Corticosterone
11-Deoxycortisol
Androstenedione
1) Describe the
biosynthesis
of the adrenal
steroid hormones
(glucocorticoids, mineralocorticoids, and androgens) and the key structural features that distinguish each class. [ojvBioSynAStrd]

2) Diagram and explain the
effects
of 21-hydrozylase deficiency (
CYP21A2
) or 17α-hydroxylase deficiency
(CYP17)
in Congenital Adrenal Hyperplasia (
CAH
). [ojvCyp21inCAH]

3)Diagram and explain the importance of p450scc (
CYP11a
) and
StAR
in steroid synthesis. [ojvC11StARssn]
Objectives
http://online.vitalsource.com/books/978-1-4698-4865-5/outline/21.7
Lipincott Key steroid hormones
Objectives NOT covered - not synthesis

Identify the statins as the main therapeutic intervention in dyslipidemia/atherosclerosis and interpret their action in terms of the inhibition of HMG CoA reductase.
For the drugs ATORVASTATIN, FLUVASTATIN, LOVASTATIN, PRAVASTATIN, ROSUVASTATIN, and SIMVASTATIN as they pertain to hyperlipidemias - describe the mechanism(s) of action, use(s), adverse effect(s), contraindication(s), and any relevant pharmacodynamic(s).
Describe the actions of each drug class on serum lipids, and compare and contrast the mechanism of each of these actions. [drug class=" BILE ACID SEQUESTRANTS, FIBRIC ACID SEQUESTRANTS, HMG CoA REDUCTASE INHIBITORS, OTHER DRUGS FOR HYPERLIPIDEMIA"].
Characterize these agents according to their action to reduce lipid synthesis or enhance removal. [drug class=" BILE ACID SEQUESTRANTS, FIBRIC ACID SEQUESTRANTS, HMG CoA REDUCTASE INHIBITORS, OTHER DRUGS FOR HYPERLIPIDEMIA"].
Describe the relevant actions of these drugs, other than on lipid metabolism (e.g. pleiotropic effects). [drug class=" BILE ACID SEQUESTRANTS, FIBRIC ACID SEQUESTRANTS, HMG CoA REDUCTASE INHIBITORS, OTHER DRUGS FOR HYPERLIPIDEMIA"].
Discuss the role of the HMG CoA reductase inhibitors in preventing acute coronary events and stroke and as possible adjuncts in the management of dementia and other pathological disorders. Consider the potential anti-inflammatory effects of “statins” on other disease states.
For the drugs ATORVASTATIN, FLUVASTATIN, LOVASTATIN, PRAVASTATIN, ROSUVASTATIN, and SIMVASTATIN as they pertain to the Summary of Classes and Specific Cardiovascular and Respiratory Drugs for Consideration - describe the mechanism(s) of action, use(s), adverse effect(s), contraindication(s), and any relevant pharmacodynamic(s).
Define, describe, and discuss the epidemiology, pathophysiology, symptoms, signs, and typical clinical course of cholelithiasis and cholecystitis. (MK)
List the water, ionic, bile salt, and bilirubin components of bile as secreted by the liver and after modification by the gallbladder.
Describe the cellular mechanisms for the hepatic uptake, conjugation, and secretion of bile salts and bilirubin.
Describe the enterohepatic circulation, including any different handling among primary and secondary bile salts, and bile acids.

3) Interpret the effect of up-regulating or down-regulating plasma cholesterol levels on the intracelleular synthesis of cholesterol, and the transcriptional regulation of genes that are involved in cholesterol homeostasis.

4) Compare and contrast the structure and function of cholesterol and cholesterol esters.
Desmolase
(CYP11A, P450scc)
NADPH
O2
17-ß-Hydroxysteroid
dehydrogenase
Aromatase
(CYP19)
11-ß-hydroxylase (CYP11B1)
21-α-Hydroxylase
(CYP21)
# Carbons 27 21 21 19 18
21-α-Hydroxylase
(CYP21)
11-ß-hydroxylase (CYP11B1)
18-α-hydroxylase
(aldosterone synthase) (CYP11B2)
3-ß-Hydroxysteroid
dehydrogenase
(3ß-HSD)
Synthesis of Steroid Hormones
Lippincott Ref
Objectives
Cytochrome p450 (CYP)
enzymes absorb at 450nm (contains heme)
mostly terminal oxidase enzymes in electron transfer chains
in ER membrane

Congenital Adrenal Hyperplasia (CAH)
21-hydroxylase deficiency (CYP21)
Mutant CYP21 > 90% CAH cases
Vit D
RDI = http://www.iom.edu/Reports/2010/Dietary-Reference-Intakes-for-Calcium-and-Vitamin-D/~/media/Files/Report%20Files/2010/Dietary-Reference-Intakes-for-Calcium-and-Vitamin-D/calciumvitd_lg.jpg?keepThis=true&TB_iframe=true&height=726&width=800;
e.g.
secreted from corpus luteum (new each menstrual cycle),
differentiation factor for mammary glands
an androgen
♂ sex hormone (produced in testes)
an estrogen
♀ main sex hormone
produced in ovary
Luteinizing
Hormone (LH)
Angiotensin II/III
Follicle Simulating Hormone (FSH)
Adrenocorticotropic Hormone (ACTH)
}
Rate
limiting
adrenal cortex
only
zona
glomeruloas
{
cholesterol
Steroidogenic Acute Regulatory protien
CYP11A1
pregnenolone
testosterone
progesterone
CYP17A
HSD17B
ER
Mitochondria
HSD3B
11-deoxycortisol
11-deoxycorticosterone
glucocorticoids
mineralocorticoids
pregnenolone
testosterone
StAR
LHR
LH
(Leutenizing Hormone)
AC
cAMP
PKA
Glucocorticoides
Modulate metabolism of carbohydrates (e.g. cortisol)
Immunosuppressive, anti-inflammatory

Mineralocorticoids
control excretion of electrolytes (e.g. aldosterone)

Androgens
Effect sex characteristics (e.g. testosterone)

Steroid Classes
Pregnenolone
precursor for all C18, C19, C21 steroids
mad in mitochondria - then goes out to cytosol
in
adrenal
cortex
zona glomerulosa
zona fasciculata
zona reticularis
P450c17
CYP17A1
17-α-Hydroxylase
17, 20-Lyase
P450c18
CYP11B2
18-α-Hydroxylase
+
-
-
-
+
+
corticosterone
aldosterone
dehydroepiandosterone
(DHEA)
cortisol
androstenedione
.
+
-
-
-
+
+
-
-
+
.
.
+
+
+
21-Hydroxylase deficiency (21-OHD) (CYP21)
1 in 60 are carrier for mutation
CYP21 mutations = 95% of cases
gene conversion (with pseudogene)
inability to synthesize cortisol
androgen excess
mineralocorticoid deficiency (often)
gene conversion events (with pseudogene)
recombination to varying degrees
clinical severity correlates with degree of enzymatic compromise
http://www.ncbi.nlm.nih.gov/pubmed/8081391
Congenital Adrenal Hyperplasia (CAH)
11-Deoxycortisol
Progesterone
11-Deoxycorticosterone
Aldosterone
CYP21
cortisol
CYP21
testosterone
↓ cortisol
↓ maintenance of blood sugar levels
↓ control of body fluids and electrolytes
↓protection against stress
↑ AdrenoCorticoTropic Hormone (ACTH)
↑ androstenedione --> ↑testosterone
3 Types
salt-losing form (most severe)
simple-virilizing form (moderate severity)
late-onset or nonclassical form (least severe)
Treatments
supplement cortisol --> ↓ACTH
(Note - cortisol ↑2-3X during stress)
Prednisolone 3 X day
At home injectable cortisol (hydrocortisone)
Dexamethasone prenatally
25X glucocorticoid potency (vs cortisol)
minimal mineralocorticoid effect.
Chronic glucocorticoids
glucocorticoid-induced osteoporosis
Glucocorticoids (if vomitting)
https://www.hopkinschildrens.org/cah/printable.html
http://www.omim.org/entry/201910
New, M.I., and Wilson, R.C. (1999). Steroid disorders in children: Congenital adrenal hyperplasia and apparent mineralocorticoid excess. PNAS 96, 12790–12797.
Ventura, A., Brunetti, G., Colucci, S., Oranger, A., Ladisa, F., Cavallo, L., Grano, M., and Faienza, M.F. (2013). Glucocorticoid-Induced Osteoporosis in Children with 21-Hydroxylase Deficiency. Biomed Res Int 2013.
CAH treated
(dexamethasone before 9th week gestation)
CAH UNtreated

Diagnosis - ACTH stimulation test
(17-OHP = 17α-hydroxyprogesterone = main 21-OHD substrate, ↑17-OHP = bad)
Heterozygotes
CAH
Nonclassical CAH
Unaffected
Classical CAH
Treatment needed before 9 weeks gestation
glucocorticoids
mineralcorticoids
androgens
http://prezi.com/w6k4jy4uzmzx/?utm_campaign=share&utm_medium=copy
CYP21
CYP21
15%
80%
5%
17-α-Hydroxylase
(CYP17)
DHEA
(Dehydroepiandrosterone)
17-α-Hydroxypregnenolone
17, 20-Lyase
(CYP17)
3ß-HSD

Figure 18.19 Composition of the plasma lipoproteins. Note the high concentration of cholesterol and cholesteryl esters in LDL.

2. Effect of endocytosed cholesterol on cellular cholesterol homeostasis: The chylomicron remnant-, IDL-, and LDL-derived cholesterol affects cellular cholesterol content in several ways (see Figure 18.20). First, expression of the gene for HMG CoA reductase is inhibited by high cholesterol, as a result of which, de novo cholesterol synthesis decreases. Additionally, degradation of the reductase is accelerated. Second, synthesis of new LDL receptor protein is reduced by decreasing the expression of the LDL receptor gene, thus limiting further entry of LDL cholesterol into cells. [Note: Regulation of the LDL receptor gene involves a SRE and SREBP-2, as was seen in the regulation of the gene for HMG CoA reductase (see p. 222). This allows coordinate regulation of the expression of these proteins.] Third, if the cholesterol is not required immediately for some structural or synthetic purpose, it is esterified by acyl CoA:cholesterol acyltransferase (ACAT). ACAT transfers a fatty acid from a fatty acyl CoA to cholesterol, producing a cholesteryl ester that can be stored in the cell (Figure 18.21). The activity of ACAT is enhanced in the presence of increased intracellular cholesterol.
3. Uptake of chemically modified LDL by macrophage scavenger receptors: In addition to the highly specific and regulated receptor-mediated pathway for LDL uptake described above, macrophages possess high levels of scavenger receptor activity. These receptors, known as scavenger receptor class A (SR-A), can bind a broad range of ligands and mediate the endocytosis of chemically modified LDL in which the lipid components or apo B have been oxidized. Unlike the LDL receptor, the scavenger receptor is not downregulated in response to increased intracellular cholesterol. Cholesteryl esters accumulate in macrophages and cause their transformation into "foam" cells, which participate in the formation of atherosclerotic plaque (Figure 18.22).
D. Metabolism of low-density lipoproteins
LDL particles contain much less TAG than their VLDL predecessors and have a high concentration of cholesterol and cholesteryl esters (Figure 18.19).
1. Receptor-mediated endocytosis: The primary function of LDL particles is to provide cholesterol to the peripheral tissues (or return it to the liver). They do so by binding to cell surface membrane LDL receptors that recognize apo B-100 (but not apo B-48). Because these LDL receptors can also bind apo E, they are known as apo B-100/apo E receptors. A summary of the uptake and degradation of LDL particles is presented in Figure 18.20. [Note: The numbers in brackets below refer to corresponding numbers on that figure.] A similar mechanism of receptor-mediated endocytosis is used for the cellular uptake and degradation of chylomicron remnants and IDLs by the liver.
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232
[1] LDL receptors are negatively charged glycoproteins that are clustered in pits on cell membranes. The cytosolic side of the pit is coated with the protein clathrin, which stabilizes the pit.
[2] After binding, the LDL—receptor complex is taken in by endocytosis. [Note: A deficiency of functional LDL receptors causes a significant elevation in plasma LDL-C. Patients with such deficiencies have type II hyperlipidemia (familial hypercholesterolemia, or FH) and premature atherosclerosis. Autosomal-dominant hypercholesterolemia can also be caused by increased activity of a protease, proprotein convertase subtilisin/kexin type 9 (PCSK9), which promotes degradation of the receptor, and by defects in apo B-100 that reduce its binding to the receptor.]
[3] The vesicle containing LDL loses its clathrin coat and fuses with other similar vesicles, forming larger vesicles called endosomes.
[4] The pH of the endosome falls (due to the proton-pumping activity of endosomal ATPase), which allows separation of the LDL from its receptor. The receptors then migrate to one side of the endosome, whereas the LDLs stay free within the lumen of the vesicle. [Note: This structure is called CURL, the compartment for uncoupling of receptor and ligand.]
[5] The receptors can be recycled, whereas the lipoprotein remnants in the vesicle are transferred to lysosomes and degraded by lysosomal acid hydrolases, releasing free cholesterol, amino acids, fatty acids, and phospholipids. These compounds can be reutilized by the cell. [Note: Storage diseases caused by rare autosomal-recessive deficiencies in the ability to hydrolyze lysosomal cholesteryl esters (late-onset Wolman disease), or to transport free cholesterol out of the lysosome (Niemann-Pick disease, Type C) have been identified.]
Figure 18.20 Cellular uptake and degradation of low-density lipoprotein (LDL). [Note: Oversupply of cholesterol accelerates the degradation of HMG CoA reductase. It also decreases synthesis of the reductase by preventing expression of its gene as seen with the LDL receptor.] ACAT = acyl CoA:cholesterol acyltransferase; HMG CoA = hydroxymethylglutaryl coenzyme A; mRNA = messenger RNA.
E. Metabolism of high-density lipoproteins
HDLs comprise a heterogeneous family of lipoproteins with a complex metabolism that is not yet completely understood. HDL particles are formed in blood by the addition of lipid to apo A-1, an apolipoprotein made by the liver and intestine and secreted into blood. [Note: HDLs are also formed within the liver and intestine.] Apo A-1 accounts for about 70% of the apolipoproteins in HDL. HDLs perform a number of important functions, including the following.
Figure 18.21 Synthesis of intracellular cholesteryl ester by ACAT. [Note: Lecithin:cholesterol acyl transferase (LCAT) is the extracellular enzyme that esterifies cholesterol using phosphatidylcholine (lecithin) as the source of the fatty acid.] CoA = coenzyme A.
1. Apolipoprotein supply: HDL particles serve as a circulating reservoir of apo C-II (the apolipoprotein that is transferred to VLDL and chylomicrons and is an activator of LPL) and apo E (the apolipoprotein required for the receptor-mediated endocytosis of IDLs and chylomicron remnants).
2. Uptake of unesterified cholesterol: Nascent HDLs are discshaped particles containing primarily phospholipid (largely phosphatidylcholine) and apolipoproteins A, C, and E. They take up cholesterol from nonhepatic (peripheral) tissues and return it to the liver as cholesteryl esters (Figure 18.23). [Note: HDL particles are excellent acceptors of unesterified cholesterol as a result of their high concentration of phospholipids, which are important solubilizers of cholesterol.]
3. Esterification of cholesterol: When cholesterol is taken up by HDL, it is immediately esterified by the plasma enzyme lecithin:cholesterol acyltransferase (LCAT, also known as PCAT, in which "P" stands for phosphatidylcholine, the source of the fatty acid). This enzyme is synthesized and secreted by the liver. LCAT binds to nascent HDL, and is activated by apo A-I. LCAT transfers the fatty acid from carbon 2 of phosphatidylcholine to cholesterol. This produces a hydrophobic cholesteryl ester, which is sequestered in the core of the HDL, and lysophosphatidylcholine, which binds to albumin. [Note: Esterification maintains the cholesterol concentration gradient, allowing continued efflux of cholesterol to HDL.] As the discoidal nascent HDL accumulates cholesteryl esters, it first becomes a spherical, relatively cholesteryl ester—poor HDL3 and, eventually, a cholesteryl ester—rich HDL2 particle that carries these esters to the liver. CETP (see p. 231) moves some of the cholesteryl esters from HDL to VLDL in exchange for TAG, relieving product inhibition of LCAT. Because VLDLs are catabolized to LDL, the cholesteryl esters transferred by CETP are ultimately
Figure 18.22 Role of oxidized lipoproteins in plaque formation in an arterial wall. LDL = low-density lipoprotein.

4. Reverse cholesterol transport: The selective transfer of cholesterol from peripheral cells to HDL, from HDL to the liver for bile acid synthesis or disposal via the bile, and to steroidogenic cells for hormone synthesis, is a key component of cholesterol homeostasis. This process of reverse cholesterol transport is, in part, the basis for the inverse relationship seen between plasma HDL concentration and atherosclerosis and for HDL's designation as the "good" cholesterol carrier. [Note: Exercise and estrogen raise HDL levels.] Reverse cholesterol transport involves efflux of cholesterol from peripheral cells to HDL, esterification of the cholesterol by LCAT, binding of the cholesteryl ester—rich HDL (HDL2) to liver (and steroidogenic cells), the selective transfer of the cholesteryl esters into these cells, and the release of
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lipid-depleted HDL (HDL3). The efflux of cholesterol from peripheral cells is mediated, at least in part, by the transport protein ABCA1. [Note: Tangier disease is a very rare deficiency of ABCA1 and is characterized by the virtual absence of HDL particles due to degradation of lipid-poor apo A-1.] The uptake of cholesteryl esters by the liver is mediated by a cell-surface receptor, SR-B1 (scavenger receptor class B type 1) that binds HDL (see p. 234 for SR-A receptors). The HDL particle itself is not taken up. Instead, there is selective uptake of the cholesteryl ester from the HDL particle. [Note: Hepatic lipase, with its ability to degrade both TAG and phospholipids, also participates in the conversion of HDL2 to HDL3.]
Figure 18.23 Metabolism of high-density lipoprotein (HDL) particles. Apo = apolipoprotein; ABCA1 = transport protein; C = cholesterol; CE = cholesteryl ester; LCAT = lecithin:cholesterol acyltransferase; VLDL = very-low-density lipoprotein; IDL = intermediate-density lipoprotein; LDL = low-density lipoprotein; CETP = cholesteryl ester transfer protein; SR-B1 = scavenger receptor B1.

ABCA1 is an ATP-binding cassette (ABC) protein. ABC proteins use energy from ATP hydrolysis to transport materials, including lipids, in and out of cells and across intracellular compartments. In addition to Tangier disease, defects in specific ABC proteins result in sitosterolemia, X-linked adrenoleukodystrophy, respiratory distress syndrome due to decreased surfactant secretion, and cystic fibrosis.

F. Role of lipoprotein (a) in heart disease
Lipoprotein (a), or Lp(a), is a particle that, when present in large quantities in the plasma, is associated with an increased risk of coronary heart disease. Lp(a) is nearly identical in structure to an LDL particle.
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Its distinguishing feature is the presence of an additional apolipoprotein molecule, apo(a), that is covalently linked at a single site to apo B-100. Circulating levels of Lp(a) are determined primarily by genetics. However, factors such as diet may play some role, as trans fatty acids have been shown to increase Lp(a), whereas ω-3 fatty acids decrease it. [Note: Apo(a) is structurally homologous to plasminogen, the precursor of a blood protease whose target is fibrin, the main protein component of blood clots (See Chapter 34 online). It is hypothesized that elevated Lp(a) slows the breakdown of blood clots that trigger heart attacks because it competes with plasminogen for binding to fibrin. The physiologic function of Lp(a) in unknown. Niacin reduces Lp(a), as well as LDL-cholesterol and TAGs, and raises HDL.]
CHAPTER 18 Cholesterol, Lipoprotein, and Steroid Metabolism

I. OVERVIEW

Cholesterol, the characteristic steroid alcohol of animal tissues, performs a number of essential functions in the body. For example, cholesterol is a structural component of all cell membranes, modulating their fluidity, and, in specialized tissues, cholesterol is a precursor of bile acids, steroid hormones, and vitamin D. It is, therefore, critically important that the cells of the body be assured an appropriate supply of cholesterol. To meet this need, a complex series of transport, biosynthetic, and regulatory mechanisms has evolved. The liver plays a central role in
regulation
of the body's cholesterol homeostasis. For example, cholesterol enters the liver's cholesterol pool from a number of sources including dietary cholesterol as well as that synthesized de novo by extrahepatic tissues and by the liver itself. Cholesterol is eliminated from the liver as unmodified cholesterol in the bile, or it can be converted to bile salts that are secreted into the intestinal lumen. It can also serve as a component of plasma lipoproteins that carry lipids to the peripheral tissues. In humans, the balance between cholesterol influx and efflux is not precise, resulting in a gradual deposition of cholesterol in the tissues, particularly in the endothelial linings of blood vessels. This is a potentially life-threatening occurrence when the lipid deposition leads to plaque formation, causing the narrowing of blood vessels (atherosclerosis) and increased risk of cardio-, cerebro-, and peripheral vascular disease. Figure 18.1 summarizes the major sources of liver cholesterol and the routes by which cholesterol leaves the liver.
II. STRUCTURE OF CHOLESTEROL

Cholesterol is a very hydrophobic compound. It consists of four fused hydrocarbon rings (A–D) called the "steroid nucleus," and it has an eight-carbon, branched hydrocarbon chain attached to carbon 17 of the D ring. Ring A has a hydroxyl group at carbon 3, and ring B has a double bond between carbon 5 and carbon 6 (Figure 18.2).

A. Sterols
Steroids with eight to ten carbon atoms in the side chain at carbon 17 and a hydroxyl group at carbon 3 are classified as sterols.
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Cholesterol is the major sterol in animal tissues. It arises from de novo synthesis and absorption of dietary cholesterol. [Note: Intestinal uptake of cholesterol is mediated, at least in part, by the protein Niemann-Pick C1-like 1 protein (NPC1-L1), the target of the drug ezetimibe that reduces absorption of dietary cholesterol (see p. 176). Plant sterols (phytosterols), such as β-sitosterol, are poorly absorbed by humans (5% absorbed as compared to 40% for cholesterol). After entering the enterocytes, they are actively transported back into the intestinal lumen. Defects in the transporter result in the rare condition of sitosterolemia. Because some cholesterol is transported back as well, plant sterols reduce the absorption of dietary cholesterol. Daily ingestion of plant sterol esters supplied, for example, in spreads or juices, is one of a number of dietary strategies to reduce plasma cholesterol levels (see p. 363).]
B. Cholesteryl esters
Most plasma cholesterol is in an esterified form (with a fatty acid attached at carbon 3, as shown in Figure 18.2), which makes the structure even more hydrophobic than free (unesterified) cholesterol. Cholesteryl esters are not found in membranes and are normally present only in low levels in most cells. Because of their hydrophobicity, cholesterol and its esters must be transported in association with protein as a component of a lipoprotein particle (see p. 227) or be solubilized by phospholipids and bile salts in the bile (see p. 226).

III. SYNTHESIS OF CHOLESTEROL

Cholesterol is synthesized by virtually all tissues in humans, although liver, intestine, adrenal cortex, and reproductive tissues, including ovaries, testes, and placenta, make the largest contributions to the body's cholesterol pool. As with fatty acids, all the carbon atoms in cholesterol are provided by acetyl coenzyme A (CoA), and nicotinamide adenine dinucleotide phosphate (NADPH) provides the reducing equivalents. The pathway is endergonic, being driven by hydrolysis of the high-energy thioester bond of acetyl CoA and the terminal phosphate bond of adenosine triphosphate (ATP). Synthesis requires enzymes in both the cytosol and the membrane of the smooth endoplasmic reticulum (ER). The pathway is responsive to changes in cholesterol concentration, and regulatory mechanisms exist to balance the rate of cholesterol synthesis within the body against the rate of cholesterol excretion. An imbalance in this
regulation
can lead to an elevation in circulating levels of plasma cholesterol, with the potential for vascular disease.
D. Regulation of cholesterol synthesis (Figure textboc below)

HMG CoA reductase is the major control point for cholesterol biosynthesis and is subject to different kinds of metabolic control.

1. Sterol-dependent regulation of gene expression: Expression of the gene for HMG CoA reductase is controlled by the transcription factor, SREBP-2 (sterol regulatory element—binding protein-2) that binds DNA at the cis-acting sterol regulatory element (SRE) upstream of the reductase gene. SREBP-2 is an integral protein of the ER membrane, and associates with a second ER membrane protein, SCAP (SREBP cleavage—activating protein). When sterol levels in the cell are low, the SREBP—SCAP complex moves from the ER to the Golgi. In the Golgi membrane, SREBP-2 is sequentially acted upon by two proteases, which generate a soluble fragment that enters the nucleus, binds the SRE, and functions as a transcription factor. This results in increased synthesis of HMG CoA reductase and, therefore, increased cholesterol synthesis (Figure 18.6). If sterols are abundant, however, they bind SCAP at its sterol-sensing domain and induce the binding of SCAP to yet other ER membrane proteins, the insigs (insulin-induced gene [products]). This results in the retention of the SCAP—SREBP complex in the ER, thereby preventing the activation of SREBP-2, and leading to downregulation of cholesterol synthesis. [Note: SREBP-1 upregulates expression of enzymes involved in fatty acid synthesis in response to insulin (see p. 184).]
2. Sterol-accelerated enzyme degradation: The reductase itself is a sterol-sensing integral protein of the ER membrane. When sterol levels in the cell are high, the enzyme binds to insig proteins. Binding leads to ubiquitination and proteasomal degradation of the reductase (see p. 247).
3. Sterol-independent phosphorylation/dephosphorylation: HMG CoA reductase activity is controlled covalently through the actions of adenosine monophosphate (AMP)-activated protein kinase ([AMPK] see p. 183) and a phosphoprotein phosphatase (see Figure 18.6). The phosphorylated form of the enzyme is inactive, whereas the dephosphorylated form is active. [Note: Because AMPK is activated by AMP, cholesterol synthesis, like fatty acid synthesis, is decreased when ATP availability is decreased.]
4. Hormonal regulation: The amount of HMG CoA reductase is controlled hormonally. An increase in insulin and thyroxine favors upregulation of the expression of the gene for the reductase. Glucagon and the glucocorticoids have the opposite effect.

Figure 18.6 Regulation of hydroxymethylglutaryl coenzyme A (HMG CoA) reductase. SRE = sterol regulatory element; SREBP = sterol regulatory element—binding protein; SCAP = SREBP cleavage—activating protein; AMPK = adenosine monophosphate—activated protein kinase; ADP = adenosine diphosphate; = phosphate; mRNA = messenger RNA.
5. Inhibition by drugs: The statin drugs (atorvastatin, fluvastatin, lovastatin, pravastatin, rosuvastatin, and simvastatin) are structural analogs of HMG CoA, and are (or are metabolized to) reversible, competitive inhibitors of HMG CoA reductase (Figure 18.7). They are used to decrease plasma cholesterol levels in patients with hypercholesterolemia.
IV. DEGRADATION OF CHOLESTEROL

The ring structure of cholesterol cannot be metabolized to CO2 and H2O in humans. Rather, the intact sterol nucleus is eliminated from the body by conversion to bile acids and bile salts, a small percentage of which is excreted in the feces, and by secretion of cholesterol into the bile, which transports it to the intestine for elimination. Some of the cholesterol in the intestine is modified by bacteria before excretion. The primary compounds made are the isomers coprostanol and cholestanol, which are reduced derivatives of cholesterol. Together with cholesterol, these compounds make up the bulk of neutral fecal sterols.
Figure 18.8 Bile acids. [Note: The ionized forms are bile salts.]

V. BILE ACIDS AND BILE SALTS

Bile consists of a watery mixture of organic and inorganic compounds. Phosphatidylcholine, or lecithin (see p. 202), and conjugated bile salts are quantitatively the most important organic components of bile. Bile can either pass directly from the liver, where it is synthesized into the duodenum through the common bile duct, or be stored in the gallbladder when not immediately needed for digestion.

A. Structure of the bile acids
The bile acids contain 24 carbons, with two or three hydroxyl groups and a side chain that terminates in a carboxyl group. The carboxyl group has a pKa of about 6. In the duodenum (pH approximately 6), this group will be protonated in half of the molecules (the bile acids) and deprotonated in the rest (the bile salts). The terms "bile acid" and "bile salt" are frequently used interchangeably, however. Both forms have hydroxyl groups that are α in orientation (they lie "below" the plane of the rings) and the methyl groups that are β (they lie "above" the plane of the rings). Therefore, the molecules have both a polar and a nonpolar face and can act as emulsifying agents in the intestine, helping prepare dietary triacylglycerol and other complex lipids for degradation by pancreatic digestive enzymes.
B. Synthesis of bile acids
Bile acids are synthesized in the liver by a multistep, multiorganelle pathway in which hydroxyl groups are inserted at specific positions on the steroid structure; the double bond of the cholesterol B ring is reduced; and the hydrocarbon chain is shortened by three carbons, introducing a carboxyl group at the end of the chain. The most common resulting compounds, cholic acid (a triol) and chenodeoxycholic acid (a diol), as shown in Figure 18.8, are called "primary" bile acids. [Note: The rate-limiting step in bile acid synthesis is the introduction of a hydroxyl group at carbon 7 of the steroid nucleus by 7-α-hydroxylase, an ER-associated cytochrome P450 monooxygenase
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found only in liver. Expression of the enzyme is downregulated by bile acids (Figure 18.9)]
C. Synthesis of conjugated bile acids
Before the bile acids leave the liver, they are conjugated to a molecule of either glycine or taurine (an end product of cysteine metabolism) by an amide bond between the carboxyl group of the bile acid and the amino group of the added compound. These new structures include glycocholic and glycochenodeoxycholic acids and taurocholic and taurochenodeoxycholic acids (Figure 18.10). The ratio of glycine to taurine forms in the bile is approximately 3:1. Addition of glycine or taurine results in the presence of a carboxyl group with a lower pKa (from glycine) or a sulfonate group (from taurine), both of which are fully ionized (negatively charged) at the alkaline pH of bile. The conjugated, ionized bile salts are more effective detergents than the unconjugated ones because of their enhanced amphipathic nature. Therefore, only the conjugated forms are found in the bile. Individuals with genetic deficiencies in the conversion of cholesterol to bile acids are treated with exogenously supplied chenodeoxycholic acid.

Figure 18.9 Synthesis of the bile acids, cholic acid and chenodeoxycholic acid, from cholesterol.

Bile salts provide the only significant mechanism for cholesterol excretion, both as a metabolic product of cholesterol and as a solubilizer of cholesterol in bile.

D. Action of intestinal flora on bile salts
Bacteria in the intestine can deconjugate (remove glycine and taurine) bile salts. They can also remove the hydroxyl group at carbon 7, producing "secondary" bile salts such as deoxycholic acid from cholic acid and lithocholic acid from chenodeoxycholic acid (Figure 18.11).

Figure 18.10 Conjugated bile salts. Note "cholic" in the names.

E. Enterohepatic circulation
Bile salts secreted into the intestine are efficiently reabsorbed (greater than 95%) and reused. The liver actively secretes bile salts into the bile. In the intestine, they are reabsorbed in the terminal ileum via a Na+-bile salt cotransporter and returned to the blood via a separate transport system. [Note: Lithocolic acid is only poorly absorbed.] They are efficiently taken up from blood by the hepatocytes via an isoform of the cotransporter and reused. [Note: Albumin binds bile salts noncovalently and transports them through the blood as was seen with fatty acids (see p. 181).] The continuous process of secretion of bile salts into the bile, their passage through the duodenum where some are deconjugated then dehydroxylated to secondary bile salts, their uptake in the ileum, and their subsequent return to the liver as a mixture of primary and secondary forms is termed the enterohepatic circulation (see Figure 18.11). Between 15 and 30 g of bile salts are secreted from the liver into the duodenum each day, yet only about 0.5 g (less than 3%) is lost daily in the feces. Approximately 0.5 g/day is synthesized from cholesterol in the liver to replace the amount lost. Bile acid sequestrants, such as cholestyramine, bind bile salts in the gut; prevent their reabsorption;
225
226
and, so, promote their excretion. They are used in the treatment of hypercholesterolemia because the removal of bile salts relieves the inhibition on bile acid synthesis in the liver, thereby diverting additional cholesterol into that pathway. [Note: Dietary fiber also binds bile salts and increases their excretion (see p. 365).]
Figure 18.11 Enterohepatic circulation of bile salts. [Note: Primary forms are converted to secondary forms by dehydroxylation.]

F. Bile salt deficiency: cholelithiasis
The movement of cholesterol from the liver into the bile must be accompanied by the simultaneous secretion of phospholipid and bile salts. If this dual process is disrupted and more cholesterol is present than can be solubilized by the bile salts and phosphatidylcholine present, the cholesterol may precipitate in the gallbladder, leading to cholesterol gallstone disease, or cholelithiasis (Figure 18.12). This disorder is typically caused by a decrease of bile acids in the bile. Cholelithiasis also may result from increased secretion of cholesterol into bile, as seen with the use of fibrates (for example, gemfibrozil) to reduce cholesterol (and triacylglycerol) in the blood. Laparoscopic cholecystectomy (surgical removal of the gallbladder through a small incision) is currently the treatment of choice. However, for patients who are unable to undergo surgery, oral administration of chenodeoxycholic acid to supplement the body's supply of bile acids results in a gradual (months to years) dissolution of the gallstones. [Note: Cholesterol stones account for over 85% of cases of cholelithiasis, with bilirubin and mixed stones accounting for the rest].
Figure 18.12 Gallbladder with gallstones.


Objectives - Choleterol Biosynthesis
Describe
cholesterol synthesis
explaining the importance of pyrophosphates, farnesyl pyrophosphate & geranylgeranyl pyrophosphate, inhibition of HMG CoA reductase with statins, and resulting change in LDL levels. [ojvCholSynFGHL]
Discuss
cholesterol receptor mediated cellular uptake
, for both LDL and oxidized LDL. [ojvCholRcpClUptk]
Distinguish the mechanisms of
cholesterol biosynthesis regulatation
via energy availability, hormones, food intake, and pharmacological manipulation (statins).
Figure 18.7 Structural similarity of hydroxymethylglutaric acid (HMG) and pravastatin, a clinically useful cholesterol-lowering drug of the "statin" family. CoA = coenzyme A.
statins
Farnasylated Proteins
(Ras)
Geranylgeranylated Proteins
(Rho, Rac)
ketone bodies
Statin - mechanisms of action (Goodman and Goodman link)

http://accessmedicine.mhmedical.com/content.aspx?sectionid=41266238&bookid=374&jumpsectionID=41275287&Resultclick=2&q=statins
statins
Lefer, A.M., Scalia, R., and Lefer, D.J. (2001). Vascular effects of HMG CoA-reductase inhibitors (statins) unrelated to cholesterol lowering: new concepts for cardiovascular disease. Cardiovascular Research 49, 281–287.
Laufs, U., and Liao, J.K. (1998). Post-transcriptional Regulation of Endothelial Nitric Oxide Synthase mRNA Stability by Rho GTPase. J. Biol. Chem. 273, 24266–24271.
geranylgeranylpyrophosphate (GGPP)
eNOS
SRE = sterol regulatory element;
SREBP = sterol regulatory element—binding protein;
SCAP = SREBP cleavage—activating protein;
INSIGs = Insulin-induced genes
when ↓[chol]
INSIGs
HMG CoA reductase
HMG CoA reductase
HMG CoA reductase
Ub
Ub
Ub
Ub
insulin, thyroxine
glucagon, glucocorticoids
↓E
and

↑Chol


↓HMG CoA reductase
↑mevalonate
GPCR
Lipid
Anchor





GGP
Rho
(a Gα)
statins → ↑eNOS mRNA → ↑NO → ↓Gα
→ ↓NFKB (↓inflammation)
→ ↓P-selectin & ↓ICAM-1 (↓cell adhesion)

↑mevalonate
statins
NFKB
lipid components or apoB
have been oxidized
INFO NOT USED
Cholesterol Synthesis and Regulation
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http://bit.ly/steroidSynthesis-ArborPrezi
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