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Transcript
Biochemistry I Integration of Metabolism: Glucose Synthesis, Transport and Regulation PPT 16.4 Chapters 16, 21, 17, 14 Gluconeogenesis OUT Chapter 14, 16, 10 IN Dr. Ray Integration of Carbohydrate and Lipid Metabolism Pentose Phosphate Pathway (PPP) • produce • produce Liver exports glucose if low blood sugar Glycogen (polymer of glucose, energy storage) Glucose-6-P Gluconeogenesis: Glycolysis: • convert • utilizes • convert • produce Pyruvate Triacylglycerol (energy storage) Pyruvate Dehydrogenase Complex: • convert • produce Ketone Bodies Cholesterol Citric Acid Cycle • convert • produce b-oxidation: • convert Acetyl CoA • produce Oxidative Phosphorylation: • • • • oxidize reduce create phosphorylate Control of Glycolysis • Flux through the glycolytic pathway must be adjusted in response to conditions both inside and outside cell. Rate of conversion of glucose into pyruvate is regulated for cellular needs: 1) production of ATP, generated by the degradation of glucose 2) form building blocks for synthetic reactions, such as the formation of fatty acids. In metabolic pathways, enzymes catalyzing essentially irreversible reactions (decrease in free energy) are potential sites of control. Hexokinase 2 Phosphofructo kinase 3 4 5 6 7 Which glycolytic enzymes are sites of control? The ________________ reactions at steps __________ , which is near equilibrium in cellular concentrations) • In gluconeogenesis, these three irreversible steps are by-passed by other reactions, in order to overcome the energetic barrier. 1 8 9 Pyruvate kinase 10 Gluconeogenesis Focus on comparing Gluconeogenesis to Glycolysis The synthesis of glucose from non-carbohydrate precursors (such as pyruvate & lactate) is called gluconeogenesis. 1. Why is this metabolic pathway important? because the _______ depends on glucose as its primary fuel and _______________ use only glucose as a fuel RBC: do not have mitochondria, NO aerobic metabolism,ATP only from glycolysis • The daily glucose requirement of the brain in a typical adult human being is about 120 g out of the160 g of glucose needed daily by the whole body. • The amount of glucose present in body fluids is about 20 g, and that readily available from glycogen, a storage form of glucose, is approximately 190 g. Direct glucose reserves are sufficient to meet glucose needs for about a day. During a longer period of not eating, glucose must be formed from non-carbohydrate sources, through gluconeogenesis mostly in the _________ and some in ______________ http://higheredbcs.wiley.com/legacy/college/boyer/0471661791/animations/animations.htm Gluconeogenesis Energetics Gluconeogenesis - NOT a reversal of glycolysis The three exergonic (irreversible) steps of glycolysis in cell: phosphorylate (1) Hexokinase (2) Phosphofructokinase (3) Pyruvate kinase make ATP by Substrate Level Phosphorylation ATP are replaced by other favorable reactions: dephosphorylate (11) Glucose-6-phosphatase (9) Fructose-1,6-bisphosphatase happens twice GTP ATP , HCO3- PyruvateOxaloacetate Oxaloacetate PEP PEP Pyruvate (1) Pyruvate carboxylase (2) Phosphoenolpyruvate carboxykinase Make high Steps 1 &2, NO details energy PEP, use ATP, GTP Energetics of Gluconeogenesis in glycolysis (step 3) Exergonic phosphorylation catalyzed by PFK DGo’ = -14.2 kJ/mol DGo’ = -16.3 kJ/mol in gluconeogenesis (step 8) Exergonic hydrolysis catalyzed by Fructose 1,6-bisphosphatase Both kinase & phosphatase reactions are exergonic. Kinase uses ATP for phosphate source and for energy source . What is the phosphoryl transfer potential of Pi (inorganic phosphate? Phosphatases hydrolyze phosphorylated alcohols (ROP), which have some small amount of free energy of hydrolysis. Non-Carbohydrate Precursors of Gluconeogenesis • Lactate – formed by lactate dehydrogenase in active skeletal muscle when rate of glycolysis exceed rate of oxidative metabolism (because of insufficient levels of oxygen). • Amino acids – from proteins in the diet and during starvation from breakdown of proteins in skeletal muscle. • Glycerol – from hydrolysis of triacylglycerols in fat cells. Glycerol can enter the glycolytic or gluconeogenic pathways through DHAP: Phosphorylate at C3 Oxidize at C2 The Cori Cycle (Fig 16.33) CORI CYCLE - Lactate formed by active muscle travels through the blood and is converted into glucose by the liver. This cycle shifts part of the metabolic burden of active muscle to the liver. Anaerobic glycolysis in muscle • The main site of gluconegenesis is in the liver, with a small amount occurring in the kidneys. • Gluconeogenesis in these two organs helps maintain glucose levels in the blood so that the brain, red blood cells and muscles can extract sufficient glucose from blood to satisfy their energy needs. Gluconeogenesis costs 6 ATP equivalents per glucose made Gluconeogenesis in liver In the liver, lactate is oxidized to pyruvate by lactate dehydrogenase. This pyruvate undergoes gluconeogenesis to produce free glucose, which is released into the blood. The liver restores the level of glucose necessary for active muscle cells to continue anaerobic glycolysis for immediate energy needs. Reciprocal Regulation of Related Pathways Glycolysis and Gluconeogenesis are coordinated, in a tissuespecific fashion, to ensure that the glucose-dependent energy needs of ALL cells are met. In a particular cell, when one pathway is upregulated, the other is downregulated. • Glycolysis produces 2 ATP • Gluconeogenesis costs 6 ATP Overall, both pathways are highly exergonic (spontaneous)! Workbook, Chapter 16 Self-Test, Q27: 1. Which of the following statements about the Cori cycle and its physiologic consequences are true? A) B) C) D) E) It involves the synthesis of glucose in muscle. It involves release of lactate by muscle. It involves lactate synthesis in the liver. It involves ATP synthesis in muscle. It involves release of glucose by the liver. Hormone Triggered Regulation of Glycogen Metabolism Epinephrine Hormones secreted by pancreas and adrenal glands: 1) Insulin (peptide) = fed state (High blood sugar) 2) Glucagon (peptide) = fasting state (Low blood sugar) 3) Epinephrine (catecholamine) = rapid energy needed Glycogen Glucose Insulin binds to Tyrosine Kinase Receptors on both Liver & Muscle cells, causing an increase in glucose uptake & glycogen degradation. Liver Cell Epinephrine binds to G-protein Coupled Receptors (GPCR) on muscle cells, causing glycogen degradation. Glucagon only binds to liver cells, causing an increase in glycogen degradation via a cAMP cascade, so that glucose is secreted into blood. Muscle Cell 10 Regulation of Glucose Uptake by GLUT Transporters • Glucose enters a cell through a homologous set of transporters called GLUT1 to GLUT5. Transporter expression is a primary means of controlling metabolism, since the plasma membrane of each type of cell expresses a different set of transporters. • Nerve cells express high concentrations of GLUT 3 (high glucose affinity). • Muscle and adipose tissue express GLUT 4 (medium glucose affinity) • Pancreatic b-cells and liver express GLUT2 (low glucose affinity) A cell can only perform biochemical reactions on compounds which it has taken up from its environment. Secretion of Insulin by Pancreatic b-cells How does the pancreas sense blood glucose levels? GLUT2 12 • Pancreatic b-cells express GLUT2 on their plasma membrane, which has a low affinity for glucose (KM = 15-20 mM), so glucose will enter these cells only when blood glucose is plentiful (after a meal). • Metabolism of glucose to CO2 increases ATP/AMP ratio (cell’s energy charge). • ATP-sensitive K+ channel closes, which alters membrane potential. • This causes a Ca2+ ion channel to open (secondary messenger), which in turn causes cell to secrete insulin into blood. Insulin Receptor – A Regulatory Protein Signal transduction (information transfer) from exterior to interior of cells, although signal molecule is NOT permeable across plasma membrane (1) How does the insulin receptor function in the plasma membrane? Inactive State: Monomer Active State: Dimer Insulin • The insulin receptor is a dimer of two identical subunits. • The separate monomers are inactive (do not bind insulin), but they come together (dimerize) around a single insulin molecule. • The dimer is active (binds insulin and becomes functional). Regulation of Glucose Uptake by GLUT Transporters Glucose transport into muscle cells is regulated by INSULIN: • GLUT 4 is stored in intracellular vesicles • When stimulated by insulin, GLUT4 transporters move to plasma membrane • Passive glucose uptake increases because more GLUT4 transporters are present in plasma membrane • When insulin decreases, GLUT4 transporters reenter cell Regulation of Glucose Uptake: http://www.wiley.com/college/f ob/quiz/quiz15/15-21.html 1) Does GLUT4 have high or low KM for glucose since muscle cells need to uptake glucose rapidly? http://www.wiley.com/college/ fob/quiz/quiz21/21-7.html OUT IN 14 Metabolic Enzymes • What types of reactions do the following enzymes catalyze? (A) Kinase – (B) Phosphatase – (C) Mutase – (D) Glycosylase – (E) Phosphorylase – breaks a glycosidic bond via phosphorolysis, thus breaks an acetal into a hemiacetal and an RO P using Pi as the nucleophile Glycogen Breakdown Removal of glucose molecules from glycogen involves 3 enzymes: Glycogen + Pi Glucose-1-P + Glycogen (n-1 residues) - Cleaves a-1,4-glycosidic bond in (n residues) glycogen, but not by hydrolysis (1) Glycogen phosphorylase: - catalyzes phosphorolysis (cleavage of a glycosidic bond [acetal] by the nucleophile Pi instead of H2O) (2) Debranching enzyme: a-1,6-glucosidase Hydrolyzes a(16) glycosidic bonds at branch points in glycogen (3) Phosphoglucomutase: Glucose-1-P Glucose-6-P (G6P feeds into glycolysis) Glycogen Metabolism in Muscle and Liver Glycogen Breakdown Glycogen Synthesis (1) Muscle cells - Produce ATP for motion (exercise) - Store glucose as glycogen when energy is not needed (resting) (2) Liver Glucose (only in liver) catalyzed by glucose-6-phosphatase - Control blood sugar - Store glucose when blood sugar is high (feeding) - Release glucose when blood sugar is low (exercise) • Several synthesis and degradation enzymes are allosterically regulated by metabolites, so that enzyme activity is adjusted to meet current needs of the cell. • Metabolism of glycogen is tightly controlled by hormones to meet metabolic needs of entire organism. General Metabolism of Glucose in Tissues Glycolysis, Gluconeogenesis, and Glycogen metabolism are coordinated, in a tissue-specific fashion, to ensure that the glucose-dependent energy needs of all cells are met. Metabolism Regulation In multistep biochemical pathways such as glycolysis, enzymes that catalyze irreversible steps are regulated, rather than enzymes that catalyze steps at equilibrium. Often feedback inhibition or feedforward activation is used for regulation of pathways. Regulation of the Citric Acid Cycle Control of the Citric Acid Cycle: • Regulated primarily by the concentrations of ATP and NADH (final products of pathway) • Key control points are two of the reactions with the largest driving force (DG’): isocitrate dehydrogenase a-ketoglutarate dehydrogenase - If cell’s energy charge [ATP]/[AMP] is high (because excess ATP is present) then these two enzymes and the entire TCA pathway is downregulated via _______________________. Amphibolic Functions of the Citric Acid Cycle TCA is both catabolic and anabolic 1. Do anabolic reactions require free energy to occur? 2. For catabolism of Acetyl CoA, are the cellular concentrations of OAA and Acetyl CoA the same? • ______________ amounts of TCA intermediates are needed to maintain the degradative (catabolic) function of the cycle. 3. Is there NET production of OAA during the Citric Acid cycle? 4. Can TCA intermediates by used to make other macromolecules? 5. Can TCA intermediates be made from sources other than acetyl CoA? http://www.wiley.com/college/fob/quiz/quiz16/16-15.html Receptor Tyrosine Kinases: Insulin Receptor From chapter 14 (FYI) Insulin: • Activation of Tyr Kinase receptors initiates a kinase cascade. • Soluble proteins are phosphorylated which converts them from inactive to activate form. • Through the sequential interaction of many different proteins on the interior side of the plasma membrane, the cell increases its uptake of glucose. • Excess glucose not needed for immediate energy is stored as glycogen. Receptor activation by covalent modification (phosphorylation) Signal Transduction Pathways (Chapter 14) FYI • Signal transduction – the multistep process by which an extracellular signal (chemical, mechanical or electrical) is amplified and converted to a intra-cellular response. • Signal transduction leads to production of intracellular second messenger (cAMP), which leads to activation of many target proteins. Adenylate Cyclase Signaling System = epinephrine (adrenaline) = cell surface receptor = cAMP cascade activates kinases = glycogen breakdown glycolysis ATP synthesis energy to run fast • Cascade quickly amplifies effect of binding a small number of hormones into release of large number of sugar (glucose) units Activating and Deactivating G-proteins in Signal Transduction Pathways FYI ON SWITCH: activate G protein (which binds to either GTP or GDP): Hormone binding to receptor causes conformational change in receptor so that G-protein releases GDP, binds GTP, and dissociates into two active components • Dephosphorylation deactivates G-protein Function of Cyclic AMP in Metabolic Regulation Activates Protein Kinase A (Chapter 10) FYI Protein Kinase A is composed of 4 subunits: 2 R + 2 C • R is regulatory subunit • C is catalytic subunit Inactive R2C2 tetramer Pseudosubstrate sequence Two active C monomers Active C cAMP binding domains Active C cAMP • No cAMP present: inactive R2C2 tetramer, because the active sites on the C subunits are occupied by a pseudosubstrate portion of R (shown in red), which prevents access to the active site by substrates. • When cAMP binds (green squares): causes allosteric conformational change of R, removes the pseudosubstrate sequence from the active sites, and tetramer dissociates into a regulatory subunit (R2) and two catalytically active subunits (2 C). • Inhibition of catalytic subunits is relieved, so activated C is now able to bind a protein substrate (phosphorylase kinase) and phosphorylate it. Slide FYI Adenylate Cyclase Signaling System Epinephrine activate Glycogen Phosphorylase Glycogen Glucose Result: Glycogen releases Glucose-1-phosphate which enters glycolysis to generate ATP for muscle movement. Integration of Carbohydrate and Lipid Metabolism Pentose Phosphate Pathway (PPP) Liver exports glucose if low blood sugar • produce NADPH • produce ribose-5-P Glycogen (polymer of glucose, energy storage) Glucose-6-P Gluconeogenesis: Glycolysis: • convert pyruvate into glucose • utilizes 4 ATP, 2 GTP & 2 NADH • convert glucose into pyruvate • produce 2 ATP & 2 NADH Pyruvate Triacylglycerol (energy storage) Pyruvate Dehydrogenase Complex: • convert pyruvate into acetyl CoA • produce NADH Ketone Bodies b-oxidation: • convert fatty acids into acetyl CoA • produce NADH & FADH2 Acetyl CoA Cholesterol Citric Acid Cycle • convert acetyl CoA to CO2 • produce NADH, FADH2 & GTP Oxidative Phosphorylation: • • • • oxidize NADH & FADH2 reduce O2 to H2O create H+ gradient across IMM phosphorylate ADP to ATP Metabolic Enzymes • What types of reactions do the following enzymes catalyze? (A) Kinase – transfers phosphate from ATP to alcohol to make RO P (B) Phosphatase – hydrolyzes phosphate from phosphorylated alcohol, thus transfers phosphate from RO P to H2O (C) Mutase – an isomerase that moves a phosphate from one alcohol to another alcohol in the same molecule (D) Glycosylase – hydrolyzes a glycosidic bond (with specific stereochemistry), thus breaks an acetal into a hemiacetal and an alcohol (E) Phosphorylase – breaks a glycosidic bond via phosphorolysis, thus breaks an acetal into a hemiacetal and an RO P using Pi as the nucleophile