Class Note for BIOC 460 at UA
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Date Created: 02/06/15
Biochemistry 460 Lecture 32 7 Dr Tischler LIPID REGULATION Related Reading Chapter 22 621622 640642 Chapter 26 742743 in Stryer 639h edition OBJECTIVES 1 Describe the events associated with covalent modi cation of hormone sensitive lipase 2 Identify the mechanism by which activators or inhibitors of phosphodiesterase affect the activity of hormonesensitive lipase 3 List the factors that induce increase synthesis or repress decrease synthesis acetyl CoA carboxylase and fatty acid synthase 4 Describe the events associated with polymerization depolymerization of acetyl CoA carboxylase 5 Discuss the events associated with covalent modi cation of acetyl CoA carboxylase 6 Discuss the events associated with covalent modi cation of HMG CoA reductase REGULATION OF PROCESSES LEADING TO FORMATION OF ACETYL CoA Parallels ofLipagenesis and axidalian of Fatty Acids In the previous lecture you learned about both the pathways for fatty acid oxidation and for fatty acid synthesis lipogenesis These pathways are summarized in Fig 1 Note that the steps in these pathways are nearly completely opposite The mitochondrial pathway for Boxidation of palmitoyl CoA commences with an oxidation step followed by hydration followed by a second oxidation step with production of FADH2 and NADH as reduced products Thiolysis then cleaves off an acetyl CoA moiety The rst step in lipogenesis which occurs in the cytoplasm rather than the mitochondria represents a reversal of the last step in the oxidation pathway in that it consists of a condensation that incorporates a two carbon unit from malonyl CoA The opposite subsequent steps are reduction then hydration followed by a second reduction step As mentioned in the pentose pathway NADPH is used in reductive biosynthesis and lipogenesis as well as cholesterol biosynthesis represent prime examples A further parallel in the two pathways is that both involve six additional cycles of the same steps to either completely oxidize or synthesize the palmitate palmitic acid Though there is no direct regulation of the pathway for Boxidation of fatty acids when liver synthesizes fatty acids their immediate oxidation is prevented by malonyl CoA inhibiting carnitine palmitoyl transferase I which forms palmitoyl carnitine for transport into the mitochondria Fig 1 This action prevents the uptake of newly formed fatty acids by the mitochondria In starvation the lack of malonyl CoA and the availability of fatty acids in the tissue determine the rate of Boxidation Lipid Regulation 1 CoA ATP PalmitoylCoA Palmitic acid inhibited by activation palmitate y malonyl CoA CPTI Use 14 NADPH to prevent 6 more times 7 malon 1 CoA I oxidation Palmitoylcarnitine through the 1 acety1yC0A mitochondria pathway total quot transport quot39o PalmitoylCoA ButyrylCOA FAD NADP E oxidation reduction 5 FADHZ NADPH hydration H7 0 H7 0 Cytosolic dehydration process NAD NADP Mitochondrial quotW OXida on 39 process E L NADH much NADPH thmlySlS t COA condensation t alonyl CoA E CH3CH212CSC0A Acetyl CoA 5 l Acetyl CoA O 6 more times through quot39 the pathway 0quot Produce 8 Acetyl CoA 7 FADH2 7 NADH total Figure 1 Parallel comparison of synthesis and oxidation of palmitate Regulation ofLipalysis and Baxidatian of Fatty Acids BOXidation of fatty acids which are released during lipolysis of stored triacylglycerol produces acetyl CoA as a product Hormone sensitive lipase triacylglycerol lipase the ratedetermining step in lipolysis is regulated via covalent modi cation Phosphorylation of the enzyme yields its active state designated as the quotaquot form Removal of the phosphate yields the inactive state designated as the quotbquot form Fig 2 Activation is initiated either by glucagon starvation or by epinephrine acute stress following binding to their speci c receptor protein Hormone binding leads to activation of adenylyl cyclase which catalyzes the formation of cyclic AMP a quotsecond messengerquot from ATP The mechanism by which hormones activate adenylyl cyclase was discussed previously Cyclic AMP in turn activates protein kinase A cAMPdependent protein kinase that phosphorylates the hormonesensitive lipase to its active quot21quot form Do not confuse this enzyme with lipoprotein lipase that hydrolyzes triacylglycerol contained in chylomicrons and VLDL The concentration of cyclic AMP can be sustained by methylxanthines eg caffeine theophylline which inhibit phosphodiesterase that catalyzes the hydrolysis of cyclic AMP to AMP If break down of cAMP decreases then its concentration is maintained higher in the cell Glucocorticoids and growth hormone also promote lipolysis by inducing the synthesis of hormonesensitive lipase Lipid Regulation 2 The rate of lipolysis is decreased through the action of protein phosphatase which removes the phosphate from hormonesensitive lipasea thus diminishing the lipase activity Insulin activates protein phosphatase and in this way counteracts the effects of glucagon or epinephrine by promoting dephosphorylation of this enzyme Note the parallel to the regulation of glycogenolysis that is also activated in liver by glucagon or epinephrine and deactivated by the counterregulatory effects of insulin Antagonism by insulin also includes the ability to decrease the intracellular concentration of cyclic AMP leading to a slower rate of lipolysis Once fatty acids are released from triacylglycerol recall that they are transported in the blood bound to albumin and delivered to the tissues where they are needed Cell me nbr ne Triacylglycerol Fatty acid Diacylglycerol HORMON ES Epinephrine ATP Glucag 533311 I Adenylyl cyclase RECEPTORS CAMP a Insulin I D I Protein K i name A nhosnhodiesterase ATP O caffeine HSLb Pi 7 t t theophylline OH 8 chljfllon GD Insulin inactive form 1n 1tion AMP HSL hormonesensitive lipase Figure 2 Hormonal activation of triacylglycerol hormonesensitive lipase Hormone signals from epinephrine or glucagon promote mobilization of fatty acids lipolysis via production of cyclic AMP Activated protein kinase A phosphorylates HSLb to the active HSLa form REGULATION OF ACETYL CoA CARBOXYLASE AND FATTY ACID SYNTHASE InductionRepression ofAcetyl CaA Curbaxylase and Fatty Acid Synthase Table I In the fed state a major fate of acetyl CoA in the liver and adipose tissue is conversion to fatty acids via lipogenesis for eventual storage as triacylglycerols The primary source of this acetyl CoA is dietary glucose Liver glucokinase is responsible for phosphorylating glucose as it enters the cell and is induced by insulin Thus following a large dietary intake of carbohydrates insulin released from the pancreas signals the liver to make more molecules of glucokinase induction With a large uptake of glucose after a meal glucose rst replenishes liver glycogen that had been consumed during the prior period of food deprivation The excess glucose in the liver is used to produce acetyl CoA for lipogenesis thus a large carbohydrate meal can be fattening Following carbohydrate intake insulin also increases glucose transport into adipose tissue the other major site of lipogenesis From a nutritional standpoint the regulation of acetyl CoA carboxylase and fatty acid synthase depends on the dietary intake of fats and carbohydrates Table l A high carbohydrate low fat diet induces the Lipid Regulation 3 synthesis of these lipogenic enzymes so that excess dietary glucose can be converted to fatty acids for fuel storage In contrast either diets high in fats or food deprivation repress reduce the amounts of these enzymes because fatty acid synthesis is undesirable under these conditions Table 1 Longterm control by induction or repression of acetyl CoA carboxylase and fatty acid synthase Physiological condition Effect Highcarbohydrate lowfat diet 7 synthesis Highfat diet l synthesis Fasting l synthesis Regulation ofAcetfyl CaA Curbaxyluse Polymerization Depolym erization acetyl CoA car boxylase polymeric 1 ATP ECOJ ADP Pi acetyl CoA malonyl CoA ACTIVE FORM palmitoylCoA promotes 5 depolymerization Citrate PromOteS polymerization 4 Pi LOWACTIVITY I acetyl CoA OH 0P03 carboxylase monomeric ATP ADP INA CTIVE FORM 3 2 Figure 3a Regulation of acetyl CoA carboxylase by citrate and palmitoyl CoA via polymerization and depolymerization respectively Acetyl CoA carboxylase eXits in three forms 1 a phosphorylated monomer 2 a nonphosphorylated monomer and 3 a polymer comprised of 3 or more nonphosphorylated monomers The most active form is the polymer Fig 3a 1 compared to its monomeric protomeric state 2 The monomer retains a low level of activity when not phosphorylated and is inactive when phosphorylated Fig 3a 3 The polymerized state only eXists in a nonphosphorylated form Allosteric binding of citrate to the dephosphorylated monomer of acetyl CoA carboxylase causes a conformational change in the subunit that facilitates polymerization and full activation of the enzyme 4 Thus excess citrate in the cytoplasm favors lipogenesis by promoting polymerization of the enzyme to this most active form Palmitoyl CoA diminishes the rate of lipogenesis by binding allosterically to the polymer causing it to destabilize 5 Thus in the presence of abundant palmitoyl CoA such as when lipolysis is active the enzyme reverts to its monomeric state This latter effect represents quotfeedbackquot inhibition as a signal for abundant levels of longchain fatty acids Lipid Regulation 4 Covalent Modi cation acetyl CoA car boxylase glucagon or polymeric 1 epinephrine ATP HCO339 ADP Fi Via cAMP 6 acetyl Co A malonyl CoA protein kinase A ACTIVE FORM cyclic AMPactivated 7 palmitoylCoA promotes 3 depolymerization 5 Citrate promotes I polymerization 4 0 Inhiblts protem phosphatase by 0 phosphorylating it 8 P protein phosphatase insulin 9 L0 WA C T I V T Y acetyl CoA OH OPOS carboxylase monomeric 2 ATP ADP NACTIVF FORM 3 AMP protein kinase active 10 ADP High ATP 12 High AMP 11 Pi TP A AMP protein kinase inactive Figure 3b Regulation of acetyl CoA carboxylase by glucagon epinephrine insulin and AMP This scenario which incorporates the scenario from Figure 3a shows the coordinated regulation of the acetyl CoA carboxylase activity This scheme is a bit different from that presented in the textbook and presents a different theory than that posed by others As it seems to be the more predominant scheme at this time it is the one for which you will be held responsible Lipid Regulation 5 Under conditions of low blood glucose glucose is unavailable as a precursor for fatty acid synthesis Consequently lipogenesis must decrease Low blood glucose leads to the secretion of glucagon which keeps acetylCoA carboxylase inactive This effect of glucagon or epinephrine is initiated by a sequence of events via cyclic AMP Fig 3b 6 and protein kinase A Fig 3b 7 that causes the monomeric form of acetyl CoA carboxylase to remain in its phosphorylated inactive state due to phosphorylation inactivation of protein phosphatase Fig 3b 8 This mechanism of inactivation of protein phosphatrase was described in lecture 28 Figure 5 Insulin counteracts the effect of glucagon or epinephrine by activating protein phosphatase by dephosphorylation Fig 3b 9 see also lecture 28 Figure 4 Phosphorylation of acetyl CoA carboxylase is catalyzed by AMP protein kinase Fig 3b 10 When energy levels are low the synthesis of fatty acids an energyconsuming process would be unfavorable AMP serves as a signal of low energy and promotes the conversion of AMP protein kinase to its active state Fig 3b 11 In contrast under high energy conditions elevated concentration of ATP fatty acid synthesis would be favorable and this condition leads to inactivation of AMP protein kinase Fig 3b 12 A comparison of gures 2 and 3b illustrates that a single hormone signal in starvation ie glucagon controls both the mobilization and decreased storage of fat by activation of hormonesensitive lipase and by shutting off acetylCoA carboxylase respectively Similarly epinephrine mobilizes fatty acids and prevents their synthesis during the stress response and following trauma REGULATION OF HMG CoA REDUCTASE FOR CHOLESTEROL BIOSYNTHESIS Regulation ofHMG CaA Reductase Synthesis by Cholesterol We discussed previously the role of hydroxymethylglutarylCoA HlVIG CoA reductase in the synthesis of cholesterol cholesterologenesis It is the committed step in cholesterologenesis and is feedback controlled through inhibition by cholesterol Despite control of this enzyme by cholesterol and by nutritional regulation through covalent modi cation elevated blood cholesterol is a common health problem Elevated blood cholesterol poses a signi cant health risk due to the increased likelihood of the development of atherosclerosis Consequently pharmaceutical companies have developed highly effective drugs statins which lower the activity of HlVIG CoA reductase by competitively inhibiting the binding of HlVIG CoA at the substrate binding site This inhibition effectively reduces the production of cholesterol The total amount of HlVIG CoA reductase enzyme in a cell is regulated by the levels of cholesterol When the amount of cholesterol in a cell is too low then the synthesis of HlVIG CoA reductase is increased This regulation is signaled through a Sterol Regulatory Element Binding Protein SREBP that is responsible for binding to and turning on transcription of the appropriate region of DNA This produces additional messenger RNA mRNA molecules that participate in translation to form more enzyme molecules SREBP is inactive in the presence of high cholesterol because it is bound to intracellular membranes eg endoplasmic reticulum so that it is unavailable to bind to DNA Fig 4 When cholesterol levels become to low SREBP is released from the membranes enters the nucleus and binds to DNA Lipid Regulation 6 ENDOPLASMIC High NU CLEUS Cholesterol 5 E E W SREBP NUCLEUS ENDOPLASMIC Low W RETICULUM Cholesterol W synthesis of HIVIG CoA reductase mRNA for HIVIG CoA Reductase Figure 4 HIVIG CoA reductase is induced when amounts of intracellular cholesterol become too low While with high cholesterol SREBP is bound to the endoplasmic reticulum and is thus rendered ineffective You will not be tested on the mechanism in Fig 4 Hormonal Regulation of Cholesterol Biosynthesis The regulation of HIVIG CoA reductase in response to nutritional status is complex In the fasting state cholesterol synthesis is undesirable because the availability of acetyl CoA from glucose via glycolysis and pyruvate dehydrogenase is limited Since glucagon is the key hormone released in fasting it is not surprising that glucagon causes inactivation of HMG CoA reductase Similarly under stress conditions when acetyl CoA must be shunted towards energy production cholesterol biosynthesis is unwanted and is prevented by epinephrine Inhibition of HIVIG CoA reductase is initiated by activation of a kinase cascade The events begin with hormonal activation of cyclic AMPdependent protein kinase A PKA Fig 5 1 The kinase cascade involves successive phosphorylation and activation of reductase kinase kinase 2 and then reductase kinase 3 Finally reductase kinase phosphorylates HIVIGCoA reductase to inactivate the enzyme and prevent the synthesis of cholesterol 4 In the fed state when glucose is abundant insulin 5 activates protein phosphatase 6 to inhibit the kinase cascade and to activate HIVIGCoA reductase just as occurs in the scenario for regulation of acetyl CoA carboxylase Hence after a meal the synthesis of fatty acids and of cholesterol is increased in parallel making use of the excess acetyl CoA derived from carbohydrates It is for this reason that a diet consistently high in carbohydrates or fats can lead to excessive storage of fats and elevated blood cholesterol In fact dietary cholesterol plays a minimal role in problems of hypercholesterolemia Generally it is dietary saturated fats that are the biggest quotculpritsquot Lipid Regulation 7 mevalonate 2 NADP insulin 5 Tn A f G P PP 6 NACTIVEFORM HR OH 7T OPO3 HMGCoA OPO 2 NADPH 3 insulin 5 2 Fr RK actlve 4 a l l 0P0 ADP PP 4 G I 3 61 RKK active 3 Pi PKA 2 94 I ATP PP lt 39 6 Insulin 5 Via 39 HR HlVIGCoA reductase I OH PKA CAMPactivated protein kinase glueagon 0f PP protein phosphatase epinephrine RK reductase kinase RKK reductase kinase kinase Figure 5 Regulation of HlVIG CoA reductase activity by phosphorylation in the presence of epinephrine or glucagon mediated Via CAlVlP and protein kinase A or by dephosphorylation in the presence of insulin mediated Via protein phosphatase Lipid Regulation 8
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