Lipid metabolism • Degradation and biosynthesis of fatty acids • Ketone bodies Fatty acids (FA) • primary fuel molecules in the fat category • main use is for long-term energy storage • high level of energy storage: fats - 38 kJ/g, carbohydrates 16 kJ/g Primary sources of FA : - dietary triacylglycerols - triacylglycerols synthesized in the liver - triacylglycerols stored in adipocytes FA must be delivered to cells where β-oxidation occurs : β FA cell -oxidation ATP • liver • heart • skeletal muscles Fatty acid degradation - β-oxidation • FA are delivered to cells by diffusion from blood capillaries • upon entry into the cell FA are immediately activated by linking them as thioesters to CoASH • activation is coupled with FA transport through the mitochondrial membrane βββ-oxidation : two-carbon fragments successively removed from the carboxyl end of activated FA producing acetyl-CoA entry into TCA cycle ATP lokalized in the mitochondrial matrix (>18 C perixosomes) Fatty acid degradation difusion activation FA entry into the cell (cytosol) acyl-CoA transport across the inner mmitochondrial membrane (carnitine shuttle ) acyl-CoA β-oxidace acetyl-CoA TCA cycle ATP Fatty acid activation – formation of acyl–CoA: O AMP + PP ATP i O C C ~SCoA OH CoASH fatty acyl coenzyme A-synthetase Carnitine shuttle – transport of long-chain FA (>12C) into the mitochondrial matrix Carnitine-acylcarnitine translocase CAT I CAT II CAT – carnitine acyltransferase carnitine (carnitine palmitoyltransferase) FA activation and transport across inner mitochondrial membrane β-Oxidation of fatty acids – spiral pathway Dehydrogenation/ oxidation Thiolytic Hydration cleavage ET chain 2 ATP Dehydrogenation/ oxidation ET chain 3 ATP Combination of FA activation, transport into the mitochondrial matrix, β-oxidation and TCA cycle Energy yield from β-oxidation of saturated FA example: stearic acid , 18 C 9 acetyl-Coa, 8 turns of β-oxidation ATP/ unit ATP produced activation -1(-2) -1(-2) 9 acetyl CoA TCA 12 108 8 FADH 2 2 16 8 (NADH + H +) 3 24 total ATP 147 (146) ββ-oxidation-oxidation of of unsaturated unsaturated fatty fatty acids acids requiresrequires additional additional enzymes enzymes– – isomerase, isomerase, NADPH-dependentNADPH-dependent reductasereductase (2,4-dienoyl-CoA (2,4-dienoyl-CoA reductase), reductase), epimeraseepimerase O O SCoA SCoA NADPH + H + Enoyl-CoA 2,4-Dienoyl-CoA isomerase reductase NADP + O SCoA SCoA O Example : Oleic acid O C ~SCoA 3 turns of β-oxidation cis-double bond γγγ β O α C ~SCoA 3 2 1 isomerase ∆3 cis ∆2 trans O β α C ~SCoA 3 2 1 hydration dehydrogenation …. Unsaturated FA provide less energy than saturated FA – less reducing equivalents (FADH 2) are produced β-oxidation of FA with an odd number of carbon They can be handled normally until the last step where propionyl CoA is produced. Three reactions are required to convert propionyl CoA to succinyl CoA which can enter to the TCA cycle. ATP + AMP + - H HCO - PP COO O 3 i O H3CCC H3CCC propionyl-CoA H SCoA carboxylase H SCoA propionyl-CoA D-methylmalonyl-CoA methylmalonyl- CoA epimerase H H H O O -OOC CCC H3CCC methylmalonyl- SCoA - H H CoA mutase COO SCoA succinyl-CoA L-methylmalonyl-CoA KetoneKetone bodiesbodies • After degradation of a fatty acid, acetyl CoA is further oxidized in the citric acid cycle. • First step in the citric acid cycle • acetyl CoA + oxaloacetate citrate • If too much acetyl CoA is produced from β-oxidation, some is converted to ketone bodies . Ketone bodies - - O CH 3-C-CH 2-COO CH 3-CH-CH 2-COO O OH CH 3-C-CH 3 acetacetate β-hydroxybutyrate acetone synthesis = ketogenesis localization: liver , mitochondrial matrix initial substrate: acetyl-CoA function: energy source physiological conditions - myocard starvation - skeletal muscles brain Formation and utilization of ketone bodies enters into TCA cycle ATP Ketogenesis O 2 CH -C- S-CoA liver (mitochondrial matrix ) 3 acetoacetyl-CoA thiolase CoASH CoASH acetoacetyl-CoA O O O CH 3-C-S-CoA CH 3-C-CH 2-C-S-CoA O OH O HMG-CoA synthase β α CoASH - O-C-CH 2-C--CH 2-C-S-CoA 5 3 1 βββ-hydroxy-βββ-methylglutaryl-CoA (HMG-CoA ) CH 3 HMG-CoA lyase O O O CH -C-CH -C-O- CH -C-S-CoA 3 2 acetoacetate 3 CO 2 O NADH+H + - CH 3-CH-CH 2-COO acetone + CH 3-C-CH 3 NAD OH βββ-hydroxybutyrate StarvationStarvation andand ketone ketone bodiesbodies hydroxybutyrate acetoacetate 7 6 5 4 3 2 (millimolar) 1 0 Blood Concentration Blood 0 1 2 3 4 Weeks of Starvation FateFate ofof ketoneketone bodies bodies • Acetoacetate and β-hydroxybutyrate Produced in the liver, diffuse in blood to other tissues. Primarily transported to muscles and brain for use as energy sources after reconverion to acetyl CoA • Acetone Only produced in small amounts, eliminated in urine or breath . H+ acetoacetateacetoacetate acetoneacetone CO 2 Spontaneous loss of CO 2 from acetoacetate. It can be detected in the breath - acetone breath Symptom of untreated diabetes mellitus or starvation conditions. Utilization of ketone bodies in extrahepatic tissues βββ-hydroxybutyrate - CH 3-CH-CH 2-COO + NAD β-hydroxybutyrate dehydrogenase OH NADH+H + O O acetacetate - sukcinyl-CoA CH 3-C-CH 2-C-O β-ketoacyl-CoA transferase O O sukcinát acetoacetyl-CoA CH 3-C-CH 2-C-S-CoA CoASH CoASH acetoacetyl-CoA thiolase acetyl-CoA O extrahepatic tissues – energy source 2 CH 3-C-S-CoA •myokard at physiological conditions CC •skeletal muscles, brain in starvation ATP Utilization of ketone bodies produced in the liver , HEART, (BRAIN) Abnormal production of ketone bodies : starvation, I. type diabetes mellitus, alcoholism secretion of glucagon activation of hormone sensitive lipase in adipocyte increased lipolysis in adipose tissue increased entry of FA into the liver ß-oxidation high production of acetyl-CoA ketogenesis + lack of oxaloacetate ketogenesis ketosis ketoacidosis ketonemia, ketonuria Ketone bodies – excretion from the organism plasma - + CH 3-C-CH 2-COO + H plasma O acetacetate ketoacidosis - + CH 3-CH-CH 2-COO + H OH acetone β-hydroxybutyrate - acetacetate + Na + H2O acetacetate - acetone 20% + β-hydroxybutyrate + Na H O β-hydroxybutyrate 2 aceton 78% 2% • increased water loss + breath moč • loss of Na urine Biosynthesis of fatty acids initial substrate : acetyl-CoA tissues: liver adipose tissue lactating mammary gland intracellular localization: cytosol - palmitic acid (C16) ER, mitochondria - elongation (C18 - C24) ER - desaturation – palmitooleic acid (C16) oleic acid (C18) arachidonic acid (C20) Reactions of FA biosynthesis- reversal course of FA ß-oxidation ß-Oxidationβ α Biosynthesis R- CH 2 - CH 2 - CO- S - dehydrogenation hydrogenation R- CH = CH 2 - CO - S - -H O hydration + H 2O 2 dehydration R- CH(OH) - CH 2 - CO - S - dehydrogenation hydrogenation R - CO - CH 2 - CO - S - thiolytic cleavage CoASH condensation R - CO - S - + CH 3- CO - SCoA Differences between FA synthesis and degradation • Intracellular localization degradation (ß-oxidation) - mitochondrial matrix biosynthesis - cytosol • Activation of an acyl degradation (ß-oxidation ) - CoA-SH biosynthesis - ACP-SH (acyl carrier protein, prosthetic group phosphopantheteine ) • Oxidoreduction cofactors degradation (ß-oxidation) - NAD +, FAD biosynthesis - NADPH + H + • Biosynthesis growth of an FA chain catalyzed by 1 multifunctional enzyme (contains aktive sites for all reactions of the synthesis) – fatty acid synthase 1. reaction of synthesis distinct from a reversal reaction of degradation malonyl CoA + acetyl CoA × acetyl CoA + acetyl CoA donor of two-carbon unit in each turn Production of cytosolic acetyl-CoA Transport of acetyl-CoA from mitochondrial matrix to cytosol in the form of citrate mitochondrial cytosol matrix oxalacetate acetyl-CoA oxalacetate acetyl-CoA inner mitochondrial ADP + Pi membrane citrate lyase citrate synthase ATP CoA CoA citrate citrate at high concntration of mitochondrial citrate, isocitrate dehydrogenase inhibited by ATP Sources of cytosolic NADPH - pentose phosphate pathway – oxidative phase - oxidation of malate – malate dehydrogenase dekarboxylating (malic enzyme): NADP + NADPH+ H + oxalacetate malate pyruvate + CO 2 Tvorba malonyl-CoA Two-carbon units areincorporated into a growing chain in the form of malonyl-CoA acetyl CoA carboxylase - CH 3-CO~CoA OO C-CH 2 -CO~S-CoA acetyl-CoA malonyl-CoA enzyme-biotin-COO - enzyme-biotin ADP+Pi ATP + CO 2 + enzyme-biotin Acetyl CoA carboxylase - kee enzyme of biosynhesis (multienzyme complex) regulation: allosteric hormonal + citrate + insulin - palmitoyl-CoA - glucagon (product) FA biosynthesis - synthesis of palmitate by the fatty acid synthase complex source of other two-carbon units 1. two-carbon unit for chain growth Regulation of fatty acid metabolism • Supply of fatty acids - liver FA - ß-oxidation at oxalacetate - citrate - ketogenesis adipocyte FA - TAG synthesis • ATP citrate (glucose supply – after meal, inhibited isocitrate dehydrogenase in TCA cycle) – after translocation to cytosol citrate activates acetyl CoA carboxylase - FA synthesis • Malonyl-CoA - inhibits carnitine acyltransferase I – long-chain FA cannot enter into mitochondria - ß-oxidation - FA accumulate in cytosol - TAG synthesis • Palmitoyl-CoA - inhibits mitochndrial translocase for citrate – citrate do not enter into cytosol - FA synthesis Regulation of fatty acid metabolism Hormonal regulation: • Insulin - dephosphorylation of acetyl CoA carboxylase (= activation) – FA synthesis • Glukagon - phosphorylation of acetyl CoA carboxylase (= inaktivace) - FA synthesis - imcreased lipolysis in adipose tissue - entry of FA into liver – ß-oxidation • Adaptive control - changes in enzymeexpresion food supply after fasting - expression of acetyl CoA carboxylase, fatty acid synthase - FA synthesis Overview of fatty acid metabolism TAG - adipose tissue HSL chylomicrones TAG GIT Liver TAG VLDL LPL LPL TAG stored in tissues FA eicosanoids complex lipids ß-oxidace biosyntéza ketone bodies cellular membranes acetyl-CoA HMG CoA steroids + TCA cycle NADH+H , FADH 2 catabolism of carbohydrates and amino acids ATP respirarory chain.