Regulation of TAG & Fatty Acid Metabolism PDF

S.H. Ackerman, PhD Regulation of TAG & Fatty acid metabolism 1 [email protected] 4213 Scott Hall LECTURE TITLE: REGULATION OF TAG & FATTY ACID METABOLISM LEARNING OBJECTIVES 1. Explain how acetyl-CoA carboxylase is regulated by key metabolit...

S.H. Ackerman, PhD Regulation of TAG & Fatty acid metabolism 1 [email protected] 4213 Scott Hall LECTURE TITLE: REGULATION OF TAG & FATTY ACID METABOLISM LEARNING OBJECTIVES 1. Explain how acetyl-CoA carboxylase is regulated by key metabolites and hormones. 2. Explain the relevance of ketone bodies. 3. List the enzymes associated with ketone metabolism, the site(s) of synthesis, the site(s) of utilization. 4. List the effects of epinephrine/glucagon and insulin on carbohydrate and lipid metabolism. OUTLINE I. Regulation of acetyl-CoA carboxylase II. Regulation of carnitine transferase I by the malonyl-CoA product of ACC2. III. Fates of acetyl-CoA released from fatty acid oxidation. IV. Ketone bodies A. Relationship between hypoglycemia and ketone bodies. B. Ketone body synthesis in the liver. C. Ketone body utilization by extrahepatic tissues. V. Carbon flux in carbohydrate and lipid metabolism in response to blood glucose. A. Pathways stimulated by glucagon B. Pathways stimulated by insulin VI. Appendices A. Compare/contrast fatty acid synthesis and fatty acid oxidation. B. Resolving the nomenclature for the thiolase super-family of enzymes.  Regulation of TAG & Fatty acid metabolism 2 Regulation of Acetyl CoA Carboxylase Implications for both fatty acid synthesis and oxidation There are two major isoforms of acetyl-CoA carboxylase in humans, ACC1 and ACC2. ACC1 is predominant in lipogenic tissues, liver, adipose tissue, mammary gland. ACC1 is located in the cytoplasm where it produces malonyl-CoA for de novo fatty acid synthesis. ACC2 is predominant in skeletal & cardiac muscle; lower levels in hepatocytes and adipocytes. ACC2 is associated with the outer membrane of mitochondria and its action is correlated principally with regulation of long chain fatty acid oxidation; malonyl-CoA is a potent inhibitor of CAT1/CPT1 of the carnitine shuttle. ACC1/ACC2 exist in cells as one of two types of homo-oligomers: 1. Short protomers (~4 polypeptide chains), which are not catalytically active. 2. Long filaments, which is the active form of the enzyme Polymerization/Depolymerization is regulated allosterically, by metabolite availability, and hormonally, by phosphorylation. In isolation, both types of effectors are able to elicit a response. However, maximal effects are observed with pairs of effectors that act synergistically. § Phosphorylation and/or Palmitoyl-CoA binding to regulatory sites, promote depolymerization. § Dephosphorylation and/or Citrate binding to regulatory sites, promotes polymerization. Epinephrine (via the b-adrenergic receptor) activates adenylyl cyclase in liver, adipose tissue, and skeletal muscle, PP1 raising the [cAMP], which activates Protein kinase A. Glucagon (via the glucagon receptor) has similar effect in hepatocytes and adipocytes. Palmitoyl-CoA is an allosteric activator of AMP-activated protein kinase (AMPK). PKA and AMPK are each capable of phosphorylating the active ACC filament, which shifts equilibrium to the inactive protomer form. Insulin brings about activation of Protein Phosphatase 1 (PP1), which binds to, and dephosphorylates, ACC protomers, shifting the equilibrium to the active filament form.  Regulation of TAG & Fatty acid metabolism 3 The trends make sense: Allosteric control of ACC, the rate-limiting enzyme of fatty acid synthesis Under energy-poor conditions that promote catabolism, the rate-limiting enzyme of the TCA cycle, isocitrate DH, is maximally active and catalyzes an exergonic reaction that pulls citrate forward in the catabolic pathway. Citrate is not available to exit mitochondria and serve as an allosteric activator of acetyl-CoA carboxylase; ACC activity is not favored when the body is in a low energy state. Isocitrate DH is inhibited by small molecules that are correlated with a high energy state (ATP, NADH), providing conditions under which the intramitochondrial [citrate] rises to a level sufficient to favor its exit through the citrate carrier to the cytosol. Citrate stimulates ACC. Under conditions of a fatty acid deficit, palmitoyl-CoA levels are not sufficient to observe allosteric inhibition of ACC, the enzyme remains active, favoring fatty acid synthesis. Once the concentration of palmitoyl-CoA exceeds the amount that is required, its concentration increases in the cytoplasm, making it available to bind and inactivate ACC. Fatty acid synthesis is disfavored. Hormonal control of ACC Insulin release in response to hyperglycemia (e.g. postprandial) stimulates anabolic pathways that favor energy storage (e.g. synthesis of glycogen and triacylglycerol (TAG)). Insulin-mediated activation of ACC1 promotes the synthesis of new fatty acids that can be incorporated in TAG for storage. Activation of ACC2 blocks the carnitine shuttle (see below). Hypoglycemia promotes carbon flux to glucose. Hypoglycemia stimulates the release of glucagon and epinephrine. These hormones activate pathways that increase blood concentrations of glucose and ketone bodies (glycogenolysis, TAG hydrolysis, which liberates glycerol for gluconeogenesis, and fatty acids for ketone body synthesis). Epinephrine and glucagon actively inhibit ACC1/ACC2 (via phosphorylation cascade) to prevent the diversion of carbons away from glucose and ketone body formation in order to maintain maximal brain function. Malonyl-CoA inhibition of carnitine acetyltransferase I The inhibition of CAT I/CPT1 by the malonyl-CoA product of ACC2 has a tissue-specific impact. Under conditions of de novo fatty acid synthesis in liver or adipose tissue, preventing the synthesis of carnitine esters with newly synthesized palmitate avoids a futile cycle. Fatty acids are not synthesized de novo in skeletal muscle. While ACC1 is virtually absent, the gene for ACC2 is highly expressed in this tissue. The inhibition of CAT1 by the malonyl-CoA product of muscle ACC2 has been linked to facilitating glucose clearance from the blood in the postprandial state. Skeletal muscle is the major site of insulin- stimulated glucose disposal (∼80%) and the efficacy of this process is correlated with muscle capacity for glucose metabolism. Long chain fatty acid oxidation floods the mitochondrial matrix with acetyl-CoA, which inhibits pyruvate Wakil, S.J., and Abu-Elheiga, L.A. (2009) J. Lipid Research, April supplement, S138-143. dehydrogenase and interferes with glucose metabolism in muscle. ACC2-mediated inhibition of CAT1 keeps intramitochondrial acetyl-CoA low and favors glucose metabolism and uptake in muscle.  Regulation of TAG & Fatty acid metabolism 4 Fates of acetyl-CoA released from FA catabolism: § Acetyl-CoA is condensed with oxaloacetate to make citrate Citrate is oxidized completely in the TCA cycle to make energy. Citrate exits mitochondria and is cleaved by ATP citrate lyase, providing acetyl-CoA in the cytoplasm for new lipid synthesis, not just FA’s, citrate also provides the carbons required for sterol biosynthesis. § Acetyl-CoA entry to the TCA cycle regenerates 4-C organic acids (succinate, fumarate, malate, oxaloacetate) that can be used in anabolic pathways to make amino acids, heme, and glucose. § Acetyl-CoA is used by liver to synthesize ketone bodies that provide an energy source alternate to glucose for the brain under conditions of hypoglycemia. Ketone bodies Hypoglycemia-induced stimulation of gluconeogenesis shifts hepatocyte lipid metabolism toward ketone body synthesis. Recall that in response to low blood glucose, liver metabolism shifts into gluconeogenesis mode and carbon flux to glucose takes precedence over everything else. The origin and fate of pyruvate both change dramatically in an effort to conserve glucose and provide cytoplasmic OAA as the precursor to PEP. Acetyl-CoA accumulates in liver mitochondria when OAA is shunted to gluconeogenesis, and is used to synthesize ketone bodies, which provide an alternative energy source for extrahepatic tissues, most notably brain.  Regulation of TAG & Fatty acid metabolism 5 Synthesis and utilization of ketone bodies Acetoacetate synthesis 1. acetyl-CoA acetyl transferase 2. HMG-CoA synthase 3. HMG lyase b-hydroxybutyrate DH interconverts acetoacetate with its reduced version, b- hydroxybutyrate. The NAD+/NADH ratio determines which species predominates The massive production of ketone bodies can be diagnosed preliminarily by the signature odor of acetone in a person’s breath and/or urine. Ketone body utilization 1. b-hydroxybutyrate DH* 2. succinyl-CoA:acetoacetate CoA transferase (thiophorase) absent in liver 3. b-ketothiolase *The action of the dehydrogenase is needed only in cases where the carbons arrive as the reduced ketone body (b- hydroxybutyrate) to target cells. Liver makes ketone bodies, but cannot use them b/c it lacks the CoA transferase enzyme that is required to convert acetoacetate to acetoacetyl-CoA. By providing the brain with an alternative source of acetyl-CoA, ketone bodies reduce glucose utilization to only what is necessary to maintain OAA levels for the TCA cycle.  Regulation of TAG & Fatty acid metabolism 6 Metabolic pathways stimulated by glucagon (low blood [glucose]); mediated by cAMP/PKA 1. TAG mobilization from white adipose tissue: the liver oxidizes the FA and makes ketone bodies from the liberated acetyl-CoA. the liver converts the glycerol backbone to glucose via gluconeogenesis. 2. Glycogenolysis in the liver via phosphorylation/activation of phosphorylase kinase. 3. Gluconeogenesis in the liver: § via phosphorylation/inhibition of PFK-2. Leads to ↓ [F26BP], which relieves F26BP inhibition of fructose-bisphosphatase 1 (FBPase-1) and favors carbon flux toward glucose. § via phosphorylation/inhibition of PK-L, the liver-specific isoform of pyruvate kinase. Promotes PEP participation in gluconeogenesis by preventing PEP conversion to pyruvate. Metabolic pathways stimulated by insulin (high blood [glucose]) 1. TAG storage in white adipose tissue by promoting TAG uptake from very low density lipoproteins (VLDL’s), which are the vehicles for transporting newly synthesized TAG released to the blood stream from the liver. 2. Glycogenesis in the liver via phosphorylation/inactivation of glycogen synthase kinase 3 (GSK-3), which permits glycogen synthase to remain unphosphorylated and active. 3. Glycolysis in the liver via activation of PFK-2. § Leads to ↑ [F26BP], which is an allosteric inhibitor of FBPase-1 and an allosteric activator of PFK-1. § Favoring carbon flux from F6P to F16BP promotes TAG synthesis in adipose tissue b/c: o F16BP cleavage generates DHAP, which is the precursor to glycerol3P in extrahepatic tissue. o Catabolic pathway leads to citrate, which supplies the carbons for fatty acid synthesis in the cytoplasm.  Regulation of TAG & Fatty acid metabolism 7 Appendix 1. Compare/contrast fatty acid synthesis and fatty acid oxidation  Regulation of TAG & Fatty acid metabolism 8 Appendix 2. Description of the thiolase super-family of enzymes. Do not memorize this information for the exam. The “thiolase” reaction is reversible and is encountered in different contexts (b-oxidation, ketone body synthesis, and cholesterol synthesis). I put together the following to help clarify confusion caused by multiple names used for the catalyzing enzyme.