chapter 12-Gluconeogenesis 3.pdf

Dr. Asmaa Abu Obaid Dr. Asmaa Abu Obaid 1 ▪ List gluconeogenic precursor ▪ List the enzymes and intermediates involved in gluconeogenesis’ ▪ Recognize the reversible and regulated steps of gluconeogenesis ▪ Discuss how blood glucose level is regulated and which hormones are involved ▪ Describe t...

Dr. Asmaa Abu Obaid Dr. Asmaa Abu Obaid 1 ▪ List gluconeogenic precursor ▪ List the enzymes and intermediates involved in gluconeogenesis’ ▪ Recognize the reversible and regulated steps of gluconeogenesis ▪ Discuss how blood glucose level is regulated and which hormones are involved ▪ Describe the mechanism of action of each of the Insulin, glucagon and epinepherin hormones ▪ Discuss regulation of gluconeogenesis 2 Dr. Asmaa Abu Obaid  Introduction ▪ The pathway for gluconeogenesis shares some steps with glycolysis, the pathway for glucose degradation, but four reactions specific to the gluconeogenic pathway are not found in the degradation pathway. ▪ These reactions replace the metabolically irreversible reactions of glycolysis. These opposing sets of reactions are an example of separate, regulated pathways for synthesis and degradation 3 Dr. Asmaa Abu Obaid Dr. Asmaa Abu Obaid 4  Gluconeogenesis ▪ large mammals that have not eaten for 16 to 24 hours have depleted their liver glycogen reserves and need to synthesize glucose to stay alive ▪ Some mammalian tissues, primarily liver and kidney, can synthesize glucose from simple precursors such as lactate and alanine. ▪ Under fasting conditions, gluconeogenesis supplies almost all of Glucose is made in the cells of the liver, kidneys (by the body’s glucose. gluconeogenesis), and small intestine (from ▪ When exercising under anaerobic hydrolysis of carbs) and exported to the conditions, muscle converts glucose to pyruvate and lactate, which travel to bloodstream the liver and are converted to glucose 5 Dr. Asmaa Abu Obaid Comparison of gluconeogenesis and glycolysis Note that many of the intermediates and enzymes are identical. All seven of the near-equilibrium reactions of glycolysis proceed in the reverse direction during gluconeogenesis Dr. Asmaa Abu Obaid 6  Gluconeogenesis Recall that glycolysis consumes two ATP molecules and generates four, for a net yield of two ATP equivalents and two molecules of NADH. Contrast this with the synthesis of one molecule of glucose by gluconeogenesis consuming a total of six ATP equivalents and two molecules of NADH. As expected, the biosynthesis of glucose requires energy and its degradation releases energy. 2 Pyruvate + 2 NADH + 4 ATP + 2 GTP + 6 H2O + 2 H+ Glucose + 2 NAD+ + 4 ADP + 2 GDP + 6 Pi 7 Dr. Asmaa Abu Obaid  Pyruvate Carboxylase  two enzymes required for synthesis of phosphoenolpyruvate.  The two steps involve a carboxylation followed by decarboxylation  Pyruvate carboxylase catalyzes a metabolically irreversible reaction—it can be allosterically activated by acetyl CoA.  Oxaloacetate can enter the citric acid cycle or serve as a precursor for glucose biosynthesis. 8 Dr. Asmaa Abu Obaid Phosphoenolpyruvate Carboxykinase → The enzyme found in bacteria, protists, fungi, and plants uses ATP while The animal version uses GTP. → In most species, the enzyme displays no allosteric kinetic properties and has no known physiological modulators. Its activity is most often affected by controls at the level of transcription of its gene 9 Dr. Asmaa Abu Obaid  Fructose 1,6-bisphosphatase ▪ The reactions of gluconeogenesis between phosphoenolpyruvate and fructose 1,6- bisphosphate are simply the reverse of the near-equilibrium reactions of glycolysis. ▪ The next reaction in the glycolysis pathway—catalyzed by phosphofructokinase-1—is metabolically irreversible 10 Dr. Asmaa Abu Obaid  Glucose 6-phosphate ▪ Although we present glucose as the final product of gluconeogenesis, this is not true in all species. ▪ In most cases, the biosynthetic pathway ends with glucose 6- phosphate. ▪ This product is an activated form of glucose. It becomes the substrate for additional carbohydrate pathways leading to synthesis of glycogen, starch and sucrose, pentose sugars, and other hexoses. 11 Dr. Asmaa Abu Obaid  Glucose 6-phosphatase ▪ In these cells, glucose 6-phosphatase is bound to the endoplasmic reticulum with its active site in the lumen. ▪ The enzyme is part of a complex that includes a glucose 6-phosphate transporter (G6PT) and a phosphate transporter. G6PT moves glucose 6- phosphate from the cytosol to the interior of the ER where it is hydrolyzed to glucose and inorganic phosphate. ▪ Phosphate is returned to the cytosol and glucose is transported to the cell surface (and the bloodstream) via the secretory pathway. 12 Dr. Asmaa Abu Obaid  Glucose 6-phosphatase A metabolically irreversible hydrolysis reaction GLUT7 transporter conveys G6P from cytosol to enzyme on the endoplasmic reticulum membrane (liver, kidney, pancreas, small intestine) Glucose is exported from the ER to bloodstream Dr. Asmaa Abu Obaid 13  Precursors for Gluconeogenesis Any metabolite that can be converted to pyruvate or oxaloacetate can be a glucose precursor Major gluconeogenic precursors in mammals: Dr. Asmaa Abu Obaid 14  Precursors for Gluconeogenesis ▪ The major gluconeogenic precursors in mammals are lactate and most amino acids, especially alanine. ▪ Glycerol, which is produced from the hydrolysis of triacylglycerols, is also a substrate for gluconeogenesis. Glycerol enters the pathway after conversion to dihydroxyacetone phosphate. 15 Dr. Asmaa Abu Obaid Gluconeogensis from Glycerol ▪ We will talk in detail about glycerol 3 P dehydrogenase complex in future chapter 16 Dr. Asmaa Abu Obaid  Lactate The interaction of glycolysis and gluconeogenesis- Glycolysis generates large the Cori cycle amounts of lactate in active muscle Red blood cells steadily produce lactate Liver lactate dehydrogenase converts lactate to pyruvate (a substrate for gluconeogensis) Glucose produced by liver is delivered to peripheral tissues via the bloodstream 17 Dr. Asmaa Abu Obaid  Amino Acids Carbon skeletons of most amino acids are catabolized to pyruvate or citric acid cycle intermediates The glucose-alanine cycle: (1) Transamination of pyruvate yields alanine which travels to the liver (2)Transamination of alanine in the liver yields pyruvate for gluconeogenesis (3) Glucose is released to the bloodstream Dr. Asmaa Abu Obaid 18  Subcellular Locations of Gluconeogenic Enzymes Gluconeogenesis enzymes are cytosolic except: (1) Glucose 6-phosphatase (endoplasmic reticulum) (2) Pyruvate carboxylase (mitochondria) (3) PEPCK (cytosol and/or mitochondria) Dr. Asmaa Abu Obaid 19 ▪ In addition to fueling the production of ATP (via glycolysis and the citric acid cycle), glucose is also a precursor of the ribose and deoxyribose moieties of nucleotides and deoxynucleotides. ▪ The pentose phosphate pathway is responsible for the synthesis of ribose as well as the production of reducing equivalents in the form of NADPH. Glucose availability is controlled by regulating the uptake and synthesis of glucose and related molecules and by regulating the synthesis and degradation of storage polysaccharides composed of glucose residues. Glucose is stored as glycogen in bacteria and animals and as starch in plants 20 Dr. Asmaa Abu Obaid Glycogen Degradation Glucose is stored as starch and glycogen Glycogen is stored in cytosolic granules in muscle and liver cells of vertebrates Glycogenolysis - degradation of glycogen Glycogen breakdown yields G1P which can be converted to G6P for metabolism via glycolysis and the citric acid cycle Muscle glycogen appears in electron micrographs as cytosolic granules with a diameter of 10 to 40 nm, about the size of ribosomes 21 2 Dr. Asmaa Abu Obaid 1 Cleavage of a glucose residue from the nonreducing end of glycogen Glycogen Phosphorylase 22 2 Dr. Asmaa Abu Obaid 2 Glycogen phosphorylase ▪ Glycogen phosphorylase is responsible for the breakdown of glycogen to produce glucose 1- phosphate. ▪ In muscle cells, glucose 1-phosphate is converted to glucose 6-phosphate that is used in glycolysis to produce ATP. ▪ In liver cells, glucose 6-phosphate is hydrolyzed to free glucose that is secreted into the bloodstream where it can be taken up by other tissues 23 Dr. Asmaa Abu Obaid Metabolism of Glucose 1-Phosphate (G1P) Phosphoglucomutase catalyzes the conversion of G1P to glucose 6-phosphate (G6P) 24 8 Dr. Asmaa Abu Obaid  Glycogen synthesis ▪ Glycogen synthesis is a polymerization reaction where glucose units are added one at a time to a growing polysaccharide chain. ▪ This reaction is catalyzed by glycogen synthase. ▪ Many polymerization reactions are processive—the enzyme remains bound to the end of the growing chain and addition reactions are very rapid ▪ The glycogen synthase reaction is distributive—the enzyme releases the growing glycogen chain after each reaction. 25 Dr. Asmaa Abu Obaid  Glycogen Synthesis Synthesis and degradation of glycogen require separate enzymatic steps Cellular glucose converted to G6P by hexokinase Three separate enzymatic steps are required to incorporate one G6P into glycogen Glycogen synthase is the major regulatory step Dr. Asmaa Abu Obaid 26  Glycogen synthase adds glucose to the nonreducing end of glycogen Dr. Asmaa Abu Obaid 27  Regulation of Glycogen Metabolism Muscle glycogen is fuel for muscle contraction Liver glycogen is mostly converted to glucose for bloodstream transport to other tissues Both mobilization and synthesis of glycogen are regulated by hormones Insulin, glucagon and epinephrine regulate mammalian glycogen metabolism Dr. Asmaa Abu Obaid 28  Hormones Regulate Glycogen Metabolism Insulin is produced by -cells of the pancreas (high levels are associated with the fed state) Insulin increases rate of glucose transport into muscle, adipose tissue via GLUT 4 transporter Insulin stimulates glycogen synthesis in the liver Dr. Asmaa Abu Obaid 29 ▪ Effect of insulin on glycogen metabolism. ▪ Insulin simulates the phosphatase activity of phosphoprotein phosphatase-1, leading to inactivation of glycogen phosphorylase and activation of glycogen synthase. 30 Dr. Asmaa Abu Obaid  Glucagon Secreted by the  cells of the pancreas in response to low blood glucose (elevated glucagon is associated with the fasted state) Stimulates glycogen degradation to restore blood glucose to steady-state levels Only liver cells are rich in glucagon receptors and therefore respond to this hormone 31 15 Dr. Asmaa Abu Obaid  ▪ When glucagon binds to its receptor it stimulates adenylate cyclase causing an increase in cAMP that leads to activation of PKA. ▪ PKA phosphorylates glycogen synthase converting the “a” form to the inactive “b” form. This blocks glycogen synthesis. ▪ PKA also phosphorylates another kinase called phosphorylase kinase The result is an increase in the rate of degradation of glycogen 32 Dr. Asmaa Abu Obaid Epinephrine (Adrenalin) Released from the adrenal glands in response to sudden energy requirement (“fight or flight”) Stimulates the breakdown of glycogen to G1P (which is converted to G6P) Increased G6P levels increase both the rate of glycolysis in muscle and glucose release to the bloodstream from the liver Dr. Asmaa Abu Obaid 33  Effects of hormones on glycogen metabolism When blood glucose is low: epinephrine and glucagon activate protein kinase A Glycogenolysis is increased (more blood glucose) Glycogen synthesis is decreased Dr. Asmaa Abu Obaid 34 Reciprocal Regulation of Glycogen Phosphorylase and Glycogen Synthase Glycogen phosphorylase (GP) and glycogen synthase COVALENT REGULATION (GS) control glycogen metabolism in liver and muscle Active form “a” Inactive form “b” cells Glycogen phosphorylase -P -OH GP and GS are reciprocally Glycogen synthase -OH -P regulated both covalently and allosterically (when one is active the other is inactive) Covalent regulation by phosphorylation (-P) and dephosphorylation (-OH) 35 19 Dr. Asmaa Abu Obaid ALLOSTERIC REGULATION by G6P ▪ GP a (active form) - inhibited by G6P ▪ GS b (inactive form) - activated by G6P 36 20 Dr. Asmaa Abu Obaid  Activation of GS and inactivation of GP by Insulin Chapter 13 Dr. Asmaa Abu Obaid Prentice Hall c2002 37  Fig 13.9 Regulation of glycogen metabolism by glucose in the liver Chapter 13 Dr. Asmaa Abu Obaid Prentice Hall c2002 38  Protein regulation at the transcription level In the fasting state In the fed state Prolonged release of glucagon from the pancreas leads to continued elevation of intracellular cAMP, that triggers increased Insulin, abundant in the fed state, acts in transcription of the PEPCK gene in the liver opposition to glucagon at the level of the and increased synthesis of PEPCK. gene reducing the rate of synthesis of After several hours, the amount of PEPCK PEPCK rises and the rate of gluconeogenesis increases 39 Dr. Asmaa Abu Obaid Summary 40 Dr. Asmaa Abu Obaid  41 Dr. Asmaa Abu Obaid  42 Dr. Asmaa Abu Obaid  43 Dr. Asmaa Abu Obaid  44 Dr. Asmaa Abu Obaid  45 Dr. Asmaa Abu Obaid  46 Dr. Asmaa Abu Obaid