Lipid Metabolism PDF

MODULE 2 4 BIOCHEMISTRY OF FAT AND NSAIDS 5 LIPID METABOLISM 1 5 LIPID METABOLISM 2  LIPID METABOLISM — CONTENTS — I. OVERVIEW LIPID METABOLI...

MODULE 2 4 BIOCHEMISTRY OF FAT AND NSAIDS 5 LIPID METABOLISM 1 5 LIPID METABOLISM 2  LIPID METABOLISM — CONTENTS — I. OVERVIEW LIPID METABOLISM VI. METABOLISM OF KETONE BODIES What are ketone bodies? II. FATTY ACID SYNTHESIS Synthesis of ketone bodies Synthesis of palmitate Fates of ketone bodies Regulation of acetyl-CoA carboxylase Pools of HMG-CoA Energetics of fatty acid synthesis VII. METABOLISM OF CHOLESTEROL III. METABOLISM OF TRIGLYCERIDES Synthesis of cholesterol Synthesis of triacylglycerides Fates of cholesterol Regulation of triacylglyceride metabolism Regulation of cholesterol biosynthesis Hormonal regulation IV. FATTY ACID OXIDATION Ubiquitination and degradation Transport of fatty acids to mitochondria Transcriptional regulation Regulation of acyl-CoA synthetase Energetics of b-oxidation V. COORDINATED REGULATION OF FATTY ACID SYNTHESIS AND DEGRADATION High carbohydrate meal Fasting 3  5. LIPID METABOLISM – LEARNING OBJECTIVES – Discuss the allosteric and hormonal (insulin and glucagon) regulation of fatty acid synthesis and degradation Explain the hormonal (insulin and glucagon) regulation of fatty acid metabolism under fasting and after a high-carbohydrate diet Define metabolism of ketone bodies, tissue-specific sources, and distinguish between ketogenic- and ketolytic tissues Identify the enzyme in cholesterol metabolism that is a target for cholesterol-reducing drugs Outline the different levels of regulation of HMG-CoA reductase 4  I. THE THREE STAGES OF INTERMEDIATE METABOLISM PROTEINS CARBOHYDRATES LIPIDS Glycolysis Gluconeogenesis Glycogenolysis Glycogenesis Stage I amino acids hexoses glycerol pentoses + fatty acids Triglyceride deamination synthesis oxidation glycolysis β-oxidation s is n tihse tino Stage II ANABOLISM CATABOLISM 0xiadtaio syens idth pyruvate -oid acyn b ox ttdy S b- Triglycerides cholesterol Facai tty Cholesterol Fa Synthesis acetyl-CoA Ketone Synthesis Ketone bodies TCA Stage III respiratory chain ATP NH3 H2O CO2 4  LIPID METABOLISM — OVERVIEW — + GLYCEROL-P TRIGLYCERIDES FATTY ACID MITOCHONDRIA TRIGLYCERIDE SYNTHESIS FATTY ACID BIOSYNTHESIS β-OXIDATION CHOLESTEROL HMG-CoA ACETYL-COA HMG-CoA KETONE BODIES CHOLESTEROL BIOSYNTHESIS KETOGENESIS CYTOSOL 6  II. FATTY ACID SYNTHESIS + GLYCEROL-P TRIGLYCERIDES FATTY ACID MITOCHONDRIA TRIGLYCERIDE SYNTHESIS FATTY ACID BIOSYNTHESIS β-OXIDATION CHOLESTEROL HMG-CoA ACETYL-COA HMG-CoA KETONE BODIES CHOLESTEROL BIOSYNTHESIS KETOGENESIS CYTOSOL 7  II. SYNTHESIS OF FATTY ACIDS Synthesis of Palmitate (16:0). The synthesis of fatty acids occurs in cytosol and requires supply of acetyl-CoA and NADPH (pentose phosphate pathway). The major pool of acetyl-CoA occurs in mitochondria; transfer of acetyl-CoA to cytosol occurs via citrate. Once in cytosol, ATP-citrate lyase metabolizes citrate to acetyl-CoA and oxaloacetate: citrate + ATP + CoA ® acetyl-CoA + ADP + oxaloacetate CYTOSOL MITOCHONDRIA ATP citrate citrate CoA ADP + Pi CoA citrate citrate lyase synthase oxaloacetate oxaloacetate glucose acetyl-CoA acetyl-CoA pentose phosphate pathway fatty NADPH acid synthesis fatty acids 8  II. SYNTHESIS OF FATTY ACIDS Synthesis of Palmitate (16:0). The catalytic machinery for fatty acid synthesis: acetyl- CoA carboxylase and fatty acid synthase. The first step in fatty acid biosynthesis is catalyzed by acetyl-CoA carboxylase (ACC) and generates malonyl-CoA: this is the commitment, irreversible step, and control of fatty acid synthesis; the enzyme is activated by citrate (the donor of acetyl-CoA units) and inhibited by palmitoyl-CoA (the SH product). SH | acetyl-CoA | Acetyl-CoA provides all the carbons carboxylase ACP FAS for fatty acids synthesis by a sequen- megacomplex ADP tial addition of acetyl-CoA to the Pi CO2 carboxyl end of the growing chain. O ATP O || || This sequence of reactions is catalyz- CH3–C–SCoA –OOC–CH –C–SCoA 2 acetyl-CoA ed by fatty acid synthase (FAS). carboxylase citrate acyl-CoA 9  II. SYNTHESIS OF FATTY ACIDS Synthesis of Palmitate (16:0). Acetyl-CoA carboxylase (ACC) is also regulated by hormones: insulin and glucagon. The overall effect of glucagon is inhibition of the enzyme: glucagon elicits the generation of the second messenger cAMP that activates PKA, which phosphorylates acetyl-CoA carboxylase. The phosphorylated form of the enzyme is inactive. P acetyl-CoA carboxylase (phosphorylated) (inactive) activate site glucagon regulation regulation hormonal hormonal GLUCAGON PKA phosphatase 1 INSULIN (non-phosphorylated) (active) O acetyl-CoA O || carboxylase || CH3–C–SCoA –OOC–CH –C–SCoA 2 citrate acyl-CoA allosteric regulation 10  II. SYNTHESIS OF FATTY ACIDS Synthesis of Palmitate (16:0). The catalytic machinery for fatty acid synthesis: acetyl- CoA carboxylase and fatty acid synthase. Malonyl-CoA and acetyl-CoA bind to a component of fatty acid synthase (the acyl carrier protein or ACP): once malonyl-CoA is formed it condenses with a molecule of acetyl-CoA; both malonyl-CoA and acetyl-CoA units are bound to an acyl carrier protein (acetyl transferase and malonyl transferase), which is a component of the multifunctional fatty acid synthase acetyl acetyl malonyl group group group C H2—COOH C H3 C H3 | | | C =O C =O C =O | SH | SH | S SH | S | S | | | | FAS FAS FAS C H3 | C H2—COOH C =O HS-CoA | | C =O HS-CoA S –CoA | S–CoA ADP Pi acetyl-CoA carboxylase CO2 ATP C H3 | C =O | S –CoA 11  acetyl malonyl group group C H3 | C H2—COOH | C =O Synthesis of Palmitate (16:0) C =O | S The catalytic machinery for fatty acid synthesis | S | | FAS C H3   The first step is the condensation of an activated acyl group | condensation C =O decarboxylation | C CO2 (an acetyl group is the first acyl group) and two carbons | C =O | SH | derived from malonyl-CoA, with the elimination of CO2 S | from the malonyl group. The net effect is extension of the FAS acyl chain by two carbons.  The product of the first reaction, a keto-acyl derivative, is NADPH C H3 |  H– C –OH reduction reduced to an alcohol (hydroxy-acyl) by a reductase activity | C NADP+ | C =O SH S | | | within fatty acid synthase and from reducing equivalents FAS from NADPH C H3 H– C |   Elimination of H2O from the hydroxyl-acyl intermediate dehydration || C –H | C =O H2O creates a double bond; the reaction is catalyzed by the SH S | | | dehydratase activity of fatty acid synthase. FAS  The double bond is reduced by a reductase activity within C H3 | CH2  NADPH | reduction C H2 | C =O NADP+ fatty acid synthase at expense of reducing equivalents from | SH S | | NADPH FAS 12  II. SYNTHESIS OF FATTY ACIDS Energetics of fatty acid biosynthesis The only step in which ATP is consumed in fatty acid biosynthesis is the reaction catalyzed by acetyl-CoA carboxylase. For example, palmitate has 16 carbons (16:0); hence, 8 molecules of acetyl-CoA will be required to form the 16-carbon palmitate. O || CH3–C–SCoA 2 NADPH SH CO2 + ATP SH | acetyl-CoA | carboxylase palmitate ADP (16:0) HSCoA + Pi FAS O || –OOC–CH –C–SCoA 2 2 NADP+ ATP used NADPH used 8 acetyl-CoA + 7 ATP + 14 NADPH → palmitate + 7 ADP + 7 Pi + 14 NADP+ + 8 HsCoA 13  FATTY ACID SYNTHESIS + GLYCEROL-P TRIGLYCERIDES FATTY ACID TRIGLYCERIDE SYNTHESIS FATTY ACID BIOSYNTHESIS INSULIN β-OXIDATION CITRATE ACYL-COA ACC GLUCAGON FAS CHOLESTEROL HMG-CoA ACETYL-COA HMG-CoA KETONE BODIES KETOGENESIS CHOLESTEROL BIOSYNTHESIS 14  WHY METABOLISM OF CARBOHYDRATES AND LIPIDS? SIGNIFICANCE: STARVE-FEED CYCLE DIABETES OBESITY ATHEROSCLEROSIS CELL SIGNALING ALZHEIMER’S DISEASE [lactate] insulin [pyruvate] glucagon HYPOXIA b cells a cells FATTY ACID SYNTHESIS Pancreas 15  III. METABOLISM OF TRIACYLGLYCERIDES + GLYCEROL-P TRIGLYCERIDES FATTY ACID TRIGLYCERIDE SYNTHESIS FATTY ACID BIOSYNTHESIS INSULIN β-OXIDATION CITRATE ACYL-COA ACC GLUCAGON FAS CHOLESTEROL HMG-CoA ACETYL-COA HMG-CoA KETONE BODIES CHOLESTEROL BIOSYNTHESIS KETOGENESIS 16  III. METABOLISM OF TRIACYLGLYCERIDES Synthesis of triacylglycerides Triacylglycerides are a major form of storage of fatty acids in liver and adipose tissue, which represent the major pools for long-term energy storage in humans. Triacylglycerols are synthesized from fatty acyl CoA and glycerol-3P. The latter originates from glucose metabolism (i.e., the reduction of dihydroxyacetone phosphate to glycerol-3P). acyl-CoA synthase acyl-CoA acyltransferase TRIACYLGLYCEROLS lipases acyl-CoA glycerol-P glycerol synthesis degradation 17  O fatty acid || III. METABOLISM OF TRIACYLGLYCERIDES R–C–OH glucose ATP + CoASH acyl-CoA synthetase glycolysis PPi + AMP originated from Glucose O CH2OH || | fatty acyl CoA R–C–SCoA HO– C –H glycerol-P | CH2–O–Pi acyltransferase O || HSCoA CH2O–C–R | HO– C –H O | || C H2–O–Pi R–C–SCoA acyltransferase HSCoA O O || || O CH2O–C–R O CH2O–C–R || | phosphatase || | R–C–O– C –H R–C–O– C –H | | CH2–O–Pi CH2–OH phosphatidic acid Pi O diacylglycerol || R–C–SCoA acyltransferase HSCoA O || O CH2O–C–R || | R–C–O– C –H O | || triacylglycerol C H2 –O–C–R 18  III. METABOLISM OF TRIACYLGLYCERIDES Regulation of triacylglyceride metabolism. Adipose tissue contains hormone-sensitive lipase (HSL) that removes fatty acids from triacylglycerols. Fatty acids and glycerol produced by lipases in adipose tissue are released to circulating blood: fatty acids are bound to serum albumin and transported to tissues for use. Glycerol returns to the liver, where is converted into dihydroxyacetone-P and enters the glycolytic pathway. albumin albumin fatty acids albumin| albumin fatty fatty acids | acids fatty acids glycerol glycerol adipocytes Hormone-Sensitive adipocytes glucose Lipase Hormone-Sensitive glucose Lipase 19 III. METABOLISM OF TRIACYLGLYCERIDES glucagon epinephrine Regulation of triacylglyceride metabolism receptor Adipose tissue triacylgycerols are mobilized AC and transported to energy-requiring tissues. caffeine ATP theophiline Glucagon and epinephrine are lipolytic phospho- hormones that activate the formation of cAMP AMP diesterase cAMP and PKA activation, which phos- phorylates (activates) the hormone-sensitive insulin HSL lipase (HSL), thus facilitating breakdown of inactive triacyl-glycerols (TG). PKA phosphatase insulin Insulin is an antilipolytic hormone that active insulin inhibits the hormone-sensitive lipase (upon HSL P activation of the protein phosphatase 1), thus TG reducing the release not only of free fatty acids but of glycerol as well. fatty acids glycerol Adipose tissue is much more sensitive to insulin than are many other tissues, which points to adipose tissue as a major site of insulin action in vivo. 20  III. METABOLISM OF TRIACYLGLYCERIDES GLUCAGON INSULIN HSL + GLYCEROL-P TRIGLYCERIDES FATTY ACID TRIGLYCERIDE SYNTHESIS FATTY ACID BIOSYNTHESIS INSULIN β-OXIDATION CITRATE ACYL-COA ACC GLUCAGON FAS CHOLESTEROL HMG-CoA ACETYL-COA HMG-CoA KETONE BODIES CHOLESTEROL BIOSYNTHESIS KETOGENESIS 21  WHY METABOLISM OF CARBOHYDRATES AND LIPIDS? SIGNIFICANCE: STARVE-FEED CYCLE DIABETES OBESITY ATHEROSCLEROSIS CELL SIGNALING ALZHEIMER’S DISEASE [lactate] insulin [pyruvate] glucagon HYPOXIA REGULATION OF HSL b cells a cells Pancreas 22  IV. FATTY ACID OXIDATION GLUCAGON INSULIN HSL + GLYCEROL-P TRIGLYCERIDES FATTY ACID TRIGLYCERIDE SYNTHESIS FATTY ACID BIOSYNTHESIS INSULIN β-OXIDATION CITRATE ACYL-COA ACC GLUCAGON FAS CHOLESTEROL HMG-CoA ACETYL-COA HMG-CoA KETONE BODIES CHOLESTEROL BIOSYNTHESIS KETOGENESIS 23  IV. FATTY ACID OXIDATION Fatty acids are used for energy production in mitochondria through the formation of NADH and FADH2 that donate electrons to the respiratory chain, thus resulting in ATP formation. Brain tissue oxidizes fatty acids to a minimal degree if at all, whereas cardiac and skeletal muscle depend heavily on fatty acids as a major energy source. fatty acid O Fatty acid oxidation or b-oxidation || R–C–OH requires activation of fatty acids to a fatty ATP + CoASH acyl-CoA acyl-CoA. This is the same process as in synthetase PPi + AMP the activation of fatty acids for synthesis malonyl-CoA of triglycerides by the enzyme acyl-CoA O || synthetase, which is inhibited by R–C–SCoA fatty acyl CoA malonyl-CoA. 24 IV. FATTY ACID OXIDATION fatty acid malonyl-CoA acyl-CoA fatty HS-CoA Transport of fatty acid to mitochondria synthase acyl-CoA β α and b-oxidation (C16) R—CH2—CH2—CH2—C—SCoA || CPT I O The inner membrane is impermeable to acyl-CoA dehydrogenase FAD FADH acyl-carnitine CoA; transport of fatty-acyl CoA requires 2 carnitine H (C R—CH —C — C —C—SCoA carnitine palmitoyl 16) transferase-1 2 H || (CPT-I) CPT II O on the outer membrane and2 carnitine enoyl-CoA H O hydratase fatty palmitoyl transferase-2 OH (CPT-II) on the acyl-CoA HS-CoA | (C16) R—CH2—CH—CH2—C—SCoA matrix side of the inner membrane).|| β-oxidation O mitochondrial matrix b- β-hydroxyacyl-CoA NAD+ Once in thedehydrogenase C14 acetyl-CoA + FADH2 + NADH NADH C12 acetyl-CoA + FADH2 + NADH oxidation of fatty acids O entails several || C10 acetyl-CoA + FADH2 + NADH (C16) R—CH2—C—CH2—C—SCoA passes with formation of acetyl-CoA, || C8 acetyl-CoA + FADH2 + NADH O acyl-CoA FADH2, and NADH in each pass. acetyltransferase HSCoA C6 acetyl-CoA + FADH2 + NADH (thiolase) C4 acetyl-CoA + FADH2 + NADH O || (C14) R—CH2—C—SCoA + CH3—C—SCoA acetyl-CoA || 25 O  IV. FATTY ACID OXIDATION GLUCAGON INSULIN HSL + GLYCEROL-P TRIGLYCERIDES FATTY ACID TRIGLYCERIDE SYNTHESIS FATTY ACID BIOSYNTHESIS INSULIN β-OXIDATION CITRATE ACYL-COA ACC ACS GLUCAGON FAS CPTI MALONYL-COA CHOLESTEROL HMG-CoA ACETYL-COA HMG-CoA KETONE BODIES CHOLESTEROL BIOSYNTHESIS KETOGENESIS 26  V. COORDINATED REGULATION OF FATTY ACID SYNTHESIS AND DEGRADATION Regulation of fatty acid metabolism in a high carbohydrate meal b-oxidation of fatty acids is unnecessary and is therefore downregulated. A high-carbohydrate meal:  raises blood glucose level;  triggers the release of insulin;  insulin-dependent protein phosphatase 1 activates ACC (acetyl-CoA carboxylase);  ACC catalyzes the formation of malonyl-CoA;  malonyl-CoA inhibits carnitine acyltransferase I (CPT I), thus preventing fatty acid entry into the mitochondrial matrix. dietary ① high blood carbohydrates glucose P ② OMM acetyl-CoA carboxylase INSULIN (phosphorylated) (inactive) ➂ glucose protein kinase phosphatase 1 glycolysis (non-phosphorylated) (active) acetyl-CoA fatty acyl HSCoA O O HSCoA carnitine || carboxylase || CH3–C–SCoA –OOC–CH –C–SCoA 2 ➄ CPT I CPT II PDH acetyl-CoA ➃ malonyl-CoA fatty carnitine fatty acyl-CoA acyl-CoA β-oxidation fatty acids NADH CYTOPLASM MITOCHONDRIA acetyl-CoA 27  V. COORDINATED REGULATION OF FATTY ACID SYNTHESIS AND DEGRADATION Regulation of fatty acid metabolism during fasting When blood glucose levels drop between meals:  glucagon is released; cAMP-dependent PKA activation;  PKA phosphorylates and inactivates ACC;  malonyl-CoA levels fall and the inhibition of fatty acid entry into mitochondria is relieved. Fatty acids enter the mitochondrial matrix to become the major fuel. Because glucagon also triggers the mobilization of fatty acids in adipose tissue, a supply low bloodof fatty acids begins arriving in the blood. low blood glucose glucose low blood ① glucose P OMM ① acetyl-CoA P OMM GLUCAGON carboxylase acetyl-CoA GLUCAGON carboxylase (phosphorylated) ② ➃ (inactive) (phosphorylated) ② ➃ (inactive) cAMP PKA phosphatase 1 ➂ cAMP PKA phosphatase 1 ➂ (non-phosphorylated) (active) (non-phosphorylated) acetyl-CoA(active) fatty acyl HSCoA O O HSCoA carnitinefatty acyl || O carboxylase acetyl-CoA || O HSCoA HSCoA ➄ carnitine CH3–C–SCoA|| –OOC–CH –C–SCoA carboxylase 2 || ➄ x CPT I CPT II CH3–C–SCoA acetyl-CoA acetyl-CoA malonyl-CoA –OOC–CH –C–SCoA 2 malonyl-CoA fatty x CPT I CPT II fatty carnitine acyl-CoA fatty carnitine acyl-CoA fatty acyl-CoA acyl-CoA β-oxidation β-oxidation fatty acids NADH fatty acids CYTOPLASM MITOCHONDRIA acetyl-CoA NADH CYTOPLASM MITOCHONDRIA acetyl-CoA28  WHY METABOLISM OF CARBOHYDRATES AND LIPIDS? SIGNIFICANCE: STARVE-FEED CYCLE DIABETES OBESITY ATHEROSCLEROSIS CELL SIGNALING ALZHEIMER’S DISEASE [lactate] insulin [pyruvate] glucagon HYPOXIA b cells a cells Pancreas COORDINATED REGULATION OF FATTY ACID SYNTHESIS AND DEGRADATION 29  VI. METABOLISM OF KETONE BODIES GLUCAGON INSULIN HSL + GLYCEROL-P TRIGLYCERIDES FATTY ACID TRIGLYCERIDE SYNTHESIS FATTY ACID BIOSYNTHESIS INSULIN β-OXIDATION CITRATE ACYL-COA ACC ACS GLUCAGON FAS CPTI MALONYL-COA CHOLESTEROL HMG-CoA ACETYL-COA HMG-CoA KETONE BODIES CHOLESTEROL BIOSYNTHESIS KETOGENESIS  VI. METABOLISM OF KETONE BODIES What are ketone bodies? There are three types of ketone bodies: acetone, acetoacetate, and D-b-hydroxybutyrate, the three of them derived from acetyl-CoA: H O | O || || H3C—C—CH3 H3C—C—CH2—C H3C—C—CH2—C || || OH | OH | | O O OH acetone acetoacetate b-hydroxybutyrate Ketone bodies are not synthesized from the metabolism of fatty acids or as intermediates in fatty acid metabolism. Ketone bodies are always synthesized from the building unit, acetyl- CoA. 31  VI. METABOLISM OF KETONE BODIES O O CH3—C CH3—C Formation of ketone bodies (ketogenesis) from acetyl- S-CoA S-CoA CoA. Healthy, well-nourished individuals produce ketone thiolase HS-CoA O bodies at a low rate. Ketone body synthesis occurs in mito- CH3—C—CH2—C || O S-CoA acetoacetyl-CoA chondria. When acetyl-CoA accumulates (starvation or acetyl-CoA HMG-CoA synthase diabetes), thiolase catalyzes the condensation of two HS-CoA OH acetyl-CoA molecules to acetoacetyl-CoA. Addition of O | O C—CH2—C—CH2—C HO | S-CoA another acetyl-CoA to acetoacetyl-CoA –catalyzed by CH3 β-hydroxy-β-methylglutaryl-CoA (HMG-CoA) HMG-CoA synthase– yields b-hydroxy-b-methylglutaryl- HMG-CoA acetyl-CoA lyase CoA (HMG-CoA) that is also an intermediate of sterol O CH3—C—CH2—C biosynthesis, but the enzyme that forms HMG-CoA || O OH acetoacetate acetoacetate D-β-hydroxybutyrate decarbosylase dehydrogenase in that pathway is cytosolic. HMG-CoA lyase is CO2 NADH OH O NAD+ | CH3—C—CH3 CH3—C—CH2—C present only in the mitochondrial matrix. || O H | OH acetone D-β-hydroxybutyrate 32  VI. METABOLISM OF KETONE BODIES Fates of ketone bodies Ketone bodies are formed in the mitochondrial matrix in the liver; they diffuse to blood and are transported to peripheral tissues: LIVER EXTRAHEPATIC TISSUES glucose fatty acids glucose fatty acids lungs acetyl-CoA (acetone) acetyl-CoA tricarboxylic ketone ketone ketone tricarboxylic acid cycle bodies bodies bodies acid cycle KETOGENESIS KETOLYSIS urine 33  VI. METABOLISM OF KETONE BODIES OH O | CH3—C—CH2—C Fates of ketone bodies: b-hydroxybutyrate, synthesiz- | OH H D-β-hydroxybutyrate ed in liver, is transferred to extrahepatic tissues, where NAD+ it is first oxidized to acetoacetate by b-hydroxybutyr- β-hydroxybutyrate dehydrogenase NADH ate dehydrogenase; succinyl-CoA transferase (SCOT) O CH3—C—CH2—C || catalyzes the conversion of acetoacetate to aceto- O OH acetoacetate acetyl-CoA, which is -cleaved into 2 acetyl-CoA succinyl-CoA succinyl-CoA transferase molecules by the action of thiolase. (SCOT) succinate O Liver does not contain the enzymes needed to CH3—C—CH2—C || O S-CoA metabolize ketone bodies (ketolysis). Hence, liver is a acetoacetyl-CoA source of ketone bodies (ketogenesis), whereas HSCoA thiolase extrahepatic tissues (heart, skeletal muscle, brain, O O CH3—C CH3—C kidney cortex) use ketone bodies as an energy source S-CoA S-CoA acetyl-CoA acetyl-CoA (ketolysis). 34  acetyl-CoA VI. METABOLISM OF KETONE BODIES There are two pools of HMG-CoA acetoacetyl-CoA There are two pools of HMG-CoA: the mitochondrial pool is used for the synthesis β-hydroxymethylglutaryl-CoA of ketone bodies (HMG-CoA) the cytosolic pool is used for the synthesis of cholesterol mitochondria cytosol ketone bodies cholesterol 35  VII. METABOLISM OF CHOLESTEROL GLUCAGON INSULIN HSL + GLYCEROL-P TRIGLYCERIDES FATTY ACID TRIGLYCERIDE SYNTHESIS FATTY ACID BIOSYNTHESIS INSULIN β-OXIDATION CITRATE ACYL-COA ACC ACS GLUCAGON FAS CPTI MALONYL-COA CHOLESTEROL HMG-CoA ACETYL-COA HMG-CoA KETONE BODIES CHOLESTEROL BIOSYNTHESIS KETOGENESIS 36  2 CH3—C=O 2 CH3—C=O | VII. METABOLISM OF CHOLESTEROL acetyl-CoA | SCoA acetyl-CoA SCoA Synthesis of cholesterol : regulatory elements in the syn- thiolase HSCoA thiolase HSCoA thesis of cholesterol and targets for cholesterol-reducing O O || || CH3—C—CH2—C=O drugs. CH3—C—CH2—C=O | acetoacetyl-CoA | SCoA acetoacetyl-CoA SCoA Thiolase catalyzes the condensation of 2 acetyl-CoA to CH —C=O CH3—C=O | HMG-CoA 3 | SCoA acetoacetyl-CoA HMG-CoA synthaseSCoA synthase HSCoA HSCoA – HMG-CoA synthase catalyzes the condensation of a COO COO– | | CH2 third acetyl-CoA to yield b-hydroxy-b-methylglutaryl- CH2 | | H3C–C–OH H3C–C–OH | CoA (HMG-CoA) Both reactions are reversible and do | CH 2 CH2 | | C=O not commit to the synthesis of cholesterol. β-hydroxymethylglutaryl-CoA C=O | β-hydroxymethylglutaryl-CoA (HMG-CoA) | SCoA (HMG-CoA) SCoA HMG-CoA reductase, an integral membrane protein of 2 NADPH 2 HMG-CoANADPH the smooth endoplasmic reticulum, catalyzes the HMG-CoA reductase reductase 2 NADP 2 NADP+ + HSCo conversion of HMG-CoA to mevalonate (NADPH + COO HSCoA– COO– | | CH2 donate two reducing equivalents). This is the CH2 | | H3C–C–OH H3C–C–OH | committed step and is the major point of regulation on | CH 2 CH2 | | CH2OH the pathway of cholesterol. mevalonate mevalonate CH OH 2 37  3 CH3–COO– VII. METABOLISM OF CHOLESTEROL acetyl-CoA 1 Synthesis of cholesterol. Cholesterol is synthesized CH3 | from acetyl-CoA in four steps: –OOC—CH —C—CH —CH —OH 2 2 2 | OH mevalonate 1 Three acetate units (2 carbons) condense to 2 generate a six-carbon intermediate (mevalonate) CH3 O O | || || CH2=C—CH2—CH2—O—P—O—P—O– 2 Mevalonate is converted into activated isoprene | O– | O– 3 isoprene units 3 Polymerization of six 5-carbon isoprene units to form the 30-carbon linear structure of squalene squalene 4 Cyclization of squalene forms the four rings of the 4 steroid nucleus, and further redox changes, removal of methyl groups, leads to the final cholesterol product, cholesterol HO 38  VII. METABOLISM OF CHOLESTEROL Fates of cholesterol. Cholesterol biosynthesis occurs largely in the liver and is exported from this tissue in two forms: bile acids and cholesteryl esters. The former are relatively hydrophilic and aid in lipid digestion; the formation of the latter is catalyzed by acyl-CoA-cholesterol acyl transferase (ACAT) that catalyzes the transfer of an activated fatty acid (acyl-CoA) to cholesterol: ACAT O O || || HO R–C–SCoA R–C–O HS-CoA This reaction converts cholesterol into a completely hydrophobic form: cholesteryl esters are the major form of cholesterol in plasma lipoproteins: 30% is found as free cholesterol and 70% as cholesteryl esters. 39  VII. METABOLISM OF CHOLESTEROL Regulation of cholesterol biosynthesis The synthesis of cholesterol is regulated at three levels: a. Hormonal regulation of HMG-CoA reductase b. Ubiquitination and degradation of HMG-CoA reductase c. Transcriptional regulation – protein levels 40  VII. METABOLISM OF CHOLESTEROL a. Hormonal regulation of HMG-CoA reductase acetyl-CoA INSULIN In humans, cholesterol production is regulated by the intracellular cholesterol concentration phosphatase β-hydroxymethyl- glutaryl-CoA and by the hormones glucagon and insulin. Glucagon stimulates the phosphorylation (inac- HMG-CoA HMG-CoA reductase reductase tivation) of HMG-CoA reductase. Insulin P inactive active stimulates the dephosphorylation (activation) of mevalonate HMG-CoA reductase by activating the protein kinase phosphatase 1, thus favoring the synthesis of GLUCAGON cholesterol. cholesteryl cholesterol HMG-CoA reductase is allosterically inhibited esters ACAT (intracellular) by the intracellular levels of cholesterol, which also activate acyl-cholesterol-acyl-transferase LDL-cholesterol (ACAT), thus increasing the concentration of (extracellular) cholesterol esters. 41  WHY METABOLISM OF CARBOHYDRATES AND LIPIDS? SIGNIFICANCE: STARVE-FEED CYCLE DIABETES OBESITY ATHEROSCLEROSIS CELL SIGNALING ALZHEIMER’S DISEASE [lactate] insulin [pyruvate] glucagon HYPOXIA b cells a cells ENDOCRINE REGULATION OF CHOLESTEROL SYNTHESIS Pancreas 42  VII. METABOLISM OF CHOLESTEROL Toll-like receptor TNFα b. Ubiquitination and degradation of HMG-CoAIL1reductase receptor TNFα receptor Cholesterol biosynthesis is regulated by ubiquitination (Ub). HMG-CoA reductase contains a sterol-binding domain, to which cholesterol (the final product) binds and IκB triggers ubiquitination of the protein, thereby preparing it for degradation by the nemo activated α P proteasome. Another protein in the ERIKK membrane is Insig β P (Insulin-induced gene) that complex binds to HMG-CoA reductase and mediates the ubiquitination (Ub) and subsequent degradation by the proteasome. P NADP+ NADP+ sterol-binding Mevalonate domain Mevalonate cholesterol ubiquitination proteasome Ub degraded proteasomeHMG-CoA ai c HMG-CoA HMG-CoA ai c ai c m ti m ti m ti do taly reductase do taly do taly n n n NADPH ca ca ca Insig proteasome cytosol Thr295–P Thr295–P GSK3! Insig ER PGC-1" PGC-1" PGC-1" cholesterol Insig-mediated proteosomal lumen binding ATP ubiquitination degradation nuclear ADP membrane NUCLEUS 43 nucleus degraded  VII. METABOLISM OF CHOLESTEROL c. Transcriptional regulation – protein levels The membrane-bound transcription factor Sterol Regulatory Element Binding Protein (SREBP) is the principal regulator of cholesterol synthesis and uptake. In humans, SREBP is bound to SCAP (SREBP Cleavage Activating Protein) in the endo- endoplasmic reticulum (ER). ER INSIG (INSulin-Induced Gene) SREBP SCAP INSIG SREBP SCAP is a key component of homeo- static regulation by controlling both the activity of SREBP and the sterol-dependent degrada- ✂ ✂ GOLGI tion degradation of HMG-CoA reductase. target genes NUCLEUS SRE HMG-CoA Reductase LDL receptor 44  REGULATION OF CHOLESTEROL SYNTHESIS c. Transcriptional regulation ER  In the presence of cholesterol, the SREBP SCAP INSIG SREBP SCAP two proteins SREBP and SCAP are PRESENCE OF ABSENCE OF  CHOLESTEROL CHOLESTEROL retained in the ER by binding to  INSIG ✂  In the absence of cholesterol, INSIG no longer ✂ GOLGI binds to SREBP-SCAP and SREBP-SCAP trans-  locates to Golgi.  In Golgi, the transcription factor domain of SREBP (SRE) is released from the membrane by  proteolytic cleavage catalyzed by proteases. SRE target genes HMG-CoA Reductase NUCLEUS LDL receptor  SRE translocates to the nucleus and activates target genes such as HMG-CoA reductase and the LDL receptor. 45  VII. METABOLISM OF CHOLESTEROL GLUCAGON INSULIN HSL + GLYCEROL-P TRIGLYCERIDES FATTY ACID TRIGLYCERIDE SYNTHESIS FATTY ACID BIOSYNTHESIS INSULIN β-OXIDATION CITRATE ACYL-COA ACC GLUCAGON FAS HMG-COA CHOLESTEROL HMG-CoA ACETYL-COA HMG-CoA KETONE BODIES REDUCTASE CHOLESTEROL BIOSYNTHESIS KETOGENESIS INSULIN CHOLESTEROL SREBP MEVALONATE GLUCAGON UBIQUITINATION 46  5. LIPID METABOLISM GROUP PROJECT Description Insulin regulates carbohydrate and lipid metabolism by acting on protein phosphatase 1. INSULIN Steps Please review your class notes and phosphorylase a P PHOSPHATASE phosphorylase b (–) glycogenolysis glycogenolysis handouts and identify the sites of insulin active inactive action Complete the scheme below according glycogen synthase D P PHOSPHATASE glycogen synthase I glycogenesis glycogenesis to the first example (glycogenolysis) by inactive active identifying how the insulin-activated acetyl-CoA PHOSPHATASE acetyl-CoA enzyme stimulates (+) or inhibits (–) carboxylase P carboxylase fatty acid fattysynthesis acid synthesis inactive active glycogenesis, fatty acid synthesis, tri- glyceride metabolism, cholesterol syn- hormone-sensitive PHOSPHATASE hormone-sensitive triglyceride degradation triglyceride metabolism thesis, and pyruvate metabolism lipase P lipase antilipolytic active inactive Mention at least 3 metabolic pathways in which glucagon antagonizes insulin PHOSPHATASE HMG-CoA HMG-CoA cholesterol synthesis reductase P reductase cholesterol synthesis inactive active Evaluation Submit the report via Brightspace PHOSPHATASE PDH P PDH pyruvate metabolism pyruvate metabolism inactive active