* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Download (a) (b)
Lipid signaling wikipedia , lookup
Catalytic triad wikipedia , lookup
Basal metabolic rate wikipedia , lookup
NADH:ubiquinone oxidoreductase (H+-translocating) wikipedia , lookup
Nicotinamide adenine dinucleotide wikipedia , lookup
Enzyme inhibitor wikipedia , lookup
Microbial metabolism wikipedia , lookup
Metabolic network modelling wikipedia , lookup
Mitogen-activated protein kinase wikipedia , lookup
Lactate dehydrogenase wikipedia , lookup
Fatty acid synthesis wikipedia , lookup
Photosynthetic reaction centre wikipedia , lookup
Adenosine triphosphate wikipedia , lookup
Metalloprotein wikipedia , lookup
Evolution of metal ions in biological systems wikipedia , lookup
Biosynthesis wikipedia , lookup
Oxidative phosphorylation wikipedia , lookup
Amino acid synthesis wikipedia , lookup
Fatty acid metabolism wikipedia , lookup
Glyceroneogenesis wikipedia , lookup
Blood sugar level wikipedia , lookup
Biochemistry wikipedia , lookup
Chapter 6. Metabolism and its regulation Section I. Carbohydrate Metabolism 1 1. What Are the Essential Features of Glycolysis? In the glycolysis pathway, a molecule of glucose is converted in 10 enzyme-catalyzed steps to two molecules of 3-carbon pyruvate. 1930s, Most of the details of this pathway were worked out by Otto Warburg, Gustav Embden, and Otto Meyerhof (German). This pathway is often referred to as the Embden–Meyerhof Pathway (EMP). Why is glycolysis so important to organisms? For some tissues and cells, glucose is the only source of metabolic energy. In addition, the product of glycolysis---pyruvate is a versatile metabolite that can be used in several ways. 2 3 Reaction 1: Glucose Is Phosphorylated Glucokinase—The First Priming Reaction by Hexokinase or G° = -RT ln Keq Hydrolysis of ATP releases 30.5 kJ/mol, and the phosphorylation of glucose “costs” 13.8 kJ/mol. Thus, the reaction liberates 16.7 kJ/mol under standard-state conditions (1mM, 25oC) 4 The incorporation of a phosphate into glucose is important for several reasons: First, phosphorylation keeps the substrate in the cell. Moreover, rapid conversion of glucose to glucose-6-phosphate keeps the intracellular concentration of glucose low, favoring faciliated diffusion of glucose into the cell. In addition, because regulatory control can be imposed only on reactions not at equilibrium, the favorable thermodynamics of this first reaction makes it an important site for regulation. 5 Enzyme that phosphorylates glucose is hexokinase, which required Mg2+ as cofactor for this reaction. There are four isozymes of hexokinase. Type I (Km=0.03mM) is the principal form in the brain. Hexokinase in skeletal muscle is a mixture of types I and II (Km=0.3mM). [normal blood glucose level is about 4 mM ] The type IV, called glucokinase, is found predominantly in the liver and pancreas with Km=10 mM. 6 Hexokinase Binds Glucose and ATP with an Induced Fit In most organisms, hexokinase occurs in a single form: a two-lobed 50-kD monomer that resembles a clamp, with a large groove in one side. (c) Glucose The (a) open and (b) closed states of yeast hexokinase. Binding of glucose (green) induces a conformation change that closes the active site. Mammalian hexokinase I contains an Nterminal domain (top) and a C-terminal domain (bottom) joined by a long -helix. Each of these domains is similar in sequence 7 and structure to yeast hexokinase Reaction 2: Phosphoglucoisomerase Catalyzes the Isomerization of Glucose-6-Phosphate G = -2.92kJ/mol Phosphoglucoisomerase, with fructose6-P (blue) bound The phosphoglucoisomerase mechanism involves opening of the pyranose ring (step 1), proton abstraction leading to enediol formation (step 2), and proton addition to the double8bond, followed by ring closure (step 3). Reaction 3: ATP Drives a Second Phosphorylation Phosphofructokinase—The Second Priming Reaction by Phosphofructokinase (PFK) is the “valve” controlling the rate of glycolysis. In addition to its role as a substrate, ATP is also an allosteric inhibitor of this enzyme. At high [ATP], PFK behaves cooperatively and the plot of enzyme activity versus [fructose-6phosphate] is sigmoid. High [ATP] thus decreases the enzyme’s affinity for fructose-6phosphate. 9 Reaction 4: Cleavage by Fructose Bisphosphate Aldolase Creates Two 3Carbon Intermediates ΔGo' = 23.9 kJ/mol ΔG= -0.23 kJ/mol Class I aldolases, found in animals and plants, characterized by the formation of a covalent Schiff base intermediate between an active-site lysine and the carbonyl group of the substrate. Class II enzymes, in fungi and bacteria, do not form the Schiff base intermediate. Instead, a zinc ion is required for its activity. 10 (a) (b) (a) The Schiff base formed between the substrate carbonyl and an activesite lysine acts as an electron sink, increasing the acidity of the hydroxyl group and facilitating cleavage as shown. The catalytic residues in the rabbit muscle enzyme are Lys229 and Asp33. (b) In Class II aldolases, an active-site Zn2 stabilizes the enolate intermediate, leading to polarization of the substrate carbonyl group. 11 Reaction 5: Interconversion of the Triose Phosphates Only one of the two triose phosphates formed by aldolase, glyceraldehyde 3-phosphate, can be directly degraded in the subsequent steps of glycolysis. The other product, dihydroxyacetone phosphate, is rapidly and reversibly converted to glyceraldehyde 3-phosphate. 12 Reaction 6: Glyceraldehyde-3-Phosphate Dehydrogenase Creates a HighEnergy Intermediate ΔGo' = +6.3 kJ/mol ΔG = -1.29 kJ/mol Glyceraldehyde-3-phosphate is oxidized to 1,3-bisphosphoglycerate by glyceraldehyde-3-phosphate dehydrogenase. The overall reaction involves both formation of a carboxylic–phosphoric anhydride (a high-energy phosphate compound) and the reduction of NAD to NADH. 13 A mechanism for the glyceraldehyde-3-phosphate dehydrogenase reaction. Reaction of an enzyme sulfhydryl with the carbonyl carbon of glyceraldehyde-3-P forms a thiohemiacetal, which loses a hydride to NAD to become a thioester. Phosphorolysis of this thioester releases 1,314 bisphosphoglycerate. Reaction 7: Phosphoryl Transfer from 1,3-Bisphosphoglycerate to ADP substrate-level phosphorylation The phosphoglycerate kinase reaction is sufficiently exergonic to pull the G-3-P dehydrogenase reaction along. 15 Reaction 8: Phosphoglycerate Mutase Catalyzes a Phosphoryl Transfer ΔGo' = 4.4 kJ/mol ΔG = 0.83 kJ/mol Phosphoglycerate mutase is a phosphoenzyme, with a phosphoryl group covalently bound to a histidine residue at the active site. This phosphoryl group is transferred to the C-2 position of the substrate to form a transient, enzymebound 2,3-bisphosphoglycerate, which then decomposes by a second phosphoryl transfer from the C-3 position of the intermediate to the histidine residue on the enzyme.. The catalytic histidine (His183) at the active site of E. coli phosphoglycerate mutase. Note that His10 is phosphorylated. 16 Reaction 9: Dehydration by Enolase Creates PEP ΔGo' = 1.8 kJ/mol ΔG = 1.1 kJ/mol What the enolase reaction does is rearrange the substrate into a form from which more of this potential energy can be released upon hydrolysis. (a) (b) The yeast enolase dimer is asymmetric. The active site of one subunit (a) contains 2phosphoglycerate, the substrate. Also shown are a Mg2+ (blue), a Li + (purple), and His159, which participates in catalysis. The other subunit (b) binds phosphoenolpyruvate, the 17 product of the enolase reaction. Reaction 10: Pyruvate Kinase Yields More ATP Pyruvate kinase is a suitable target site for regulation of glycolysis and possesses allosteric sites for numerous effectors. It is activated by AMP and fructose-1,6-bisphosphate and inhibited by ATP, acetyl-CoA, and alanine. 18 What are The metabolic fates of NADH and Pyruvate Partially depends on the availability of oxygen. Under aerobic conditions, pyruvate can be sent into the citric acid cycle, whereas NADH is reoxidized to NAD+ in the mitochondrial electrontransport chain. Anaerobic Metabolism of Pyruvate Leads to Lactate or Ethanol Pyruvate decarboxylase (a) Pyruvate reduction to ethanol in yeast. (b) In oxygen-depleted muscle, NAD is 19 regenerated in the lactate dehydrogenase reaction. GLYCOLYSIS PATHWAY 20 3. The Tricarboxylic Acid Cycle For most eukaryotic cells and many bacteria, which live under aerobic conditions and oxidize their organic fuels to carbon dioxide and water--cellular respiration. First stage, organic fuel molecules are oxidized to yield two-carbon fragments of acetylcoenzyme A (acetyl-CoA). Second stage, the acetyl groups are oxidized to CO2 via TCA pathway; the energy released is conserved in the reduced electron carriers NADH and FADH2. Third stage, reduced coenzymes are via the respiratory chain, energy released is conserved in the form of ATP. 21 1. Production of Acetyl-CoA The pyruvate dehydrogenase complex requires five coenzymes: thiamine pyrophosphate (TPP), flavin adenine dinucleotide (FAD), coenzyme A (CoA), nicotinamide adenine dinucleotide (NAD), and lipoate. Reactive thiol group 22 The Pyruvate Dehydrogenase Complex Consists of Three Distinct Enzymes: pyruvate dehydrogenase (E1), dihydrolipoyl transacetylase (E2), and dihydrolipoyl dehydrogenase (E3)—each present in multiple copies. In the bovine enzyme complex, 60 identical copies of E2 form a dodecahedron core with a diameter of about 25 nm. E2 has three domains: the amino-terminal lipoyl domain; the central E1- and E3-binding domain; and the inner-core acyltransferase domain. The active site of E1 and E3 has bound TPP and FAD, respectively. The attachment of lipoate to E2 produces a long, flexible arm that can move from the active site of E1 to the active sites of E2 and E3. 23 In Substrate Channeling, Intermediates Never Leave the Enzyme Surface (1) Pyruvate reacts with the bound TPP of E1, undergoing decarboxylation (2) E1 catalyze the transfer of two electrons and the acetyl group from TPP to lipoyllysyl group of E2, to form the acetyl thioester of the reduced lipoyl group. (3) Transesterification in which the -SH group of CoA replaces the -SH group of E2 to yield acetyl-CoA and the fully reduced form of the lipoyl group. (4) E3 promotes transfer of two hydrogen atoms from the reduced lipoyl groups to the FAD. (5) The reduced FADH2 transfers a hydride ion to NAD, forming NADH. 2. Citric Acid Cycle 25 Step 1. Formation of Citrate In this reaction the methyl carbon of the acetyl group is joined to the carbonyl group (C-2) of oxaloacetate. Citroyl-CoA is a transient intermediate formed on the active site of the enzyme. 26 CoA-SH 3 H2O 2 27 Step 2. Formation of Isocitrate via cis-Aconitate ∆G0=13.3 kJ/mol This reaction is pulled to the right because isocitrate is rapidly consumed in the next step of the cycle, lowering its steady-state concentration. The active site of aconitase. The iron–sulfur cluster (pink) is coordinated by cysteines (orange) and isocitrate (purple) 28 Step 3. Oxidation of Isocitrate to α-Ketoglutarate and CO2 Two different forms of isocitrate dehydrogenase, one requiring NAD as electron acceptor (occurs in the mitochondrial matrix and serves in the citric acid cycle) and the other requiring NADP found in both the mitochondrial matrix and the cytosol (reductive anabolic reactions). Both the two forms need Mg2+ as cofactor. 29 Step 4. Oxidation of α-Ketoglutarate to Succinyl-CoA and CO2 In this reaction, NAD serves as electron acceptor and CoA as the carrier of the succinyl group. This reaction is virtually identical to the pyruvate dehydrogenase reaction, and the α-ketoglutarate dehydrogenase complex closely resembles the PDH complex in both structure and function (E1, E2 and E3). 30 Step5. Conversion of Succinyl-CoA to Succinate In step 1 a phosphoryl group replaces the CoA of succinyl-CoA, forming a high-energy acyl phosphate. In step 2 the succinyl phosphate donates its phosphoryl group to a His residue on the enzyme, forming a highenergy phosphohistidyl enzyme. In step 3 the phosphoryl group is transferred from the His residue to GDP (or ADP), forming GTP (or ATP). 31 Step 6. Oxidation of Succinate to Fumarate Step 7. Hydration of Fumarate to Malate This enzyme is highly stereospecific; it catalyzes hydration of the trans double bond of fumarate but not the cis double bond of. In the reverse direction (from Lmalate to fumarate), fumarase is equally stereospecific: D-malate is not a substrate. 32 Step 8. Oxidation of Malate to Oxaloacetate ∆Go = 29.7 kJ/mol In intact cells, oxaloacetate is continually removed by the highly exergonic citrate synthase reaction. This keeps the concentration of oxaloacetate in the cell extremely low (<10-6 M), pulling the malate dehydrogenase reaction toward the formation of oxaloacetate. 33 The Energy of Oxidations in the Cycle Is Efficiently Conserved The citric acid cycle directly generates only one ATP, four NADH and one FADH2. In oxidative phosphorylation, passage of two electrons from NADH (or FADH2) to O2 drives the formation of about 2.5 ATP (or about1.5 ATP). In round numbers, 32×30.5 kJ/mol = 976 kJ/mol, or 34% of the theoretical maximum of about 2,840 kJ/mol available from the complete oxidation of glucose (standard free energy changes) were conserved. When corrected for the actual free energy required to form ATP within cells, the calculated efficiency of the process is closer to 65%. 34 Citric Acid Cycle Components Are Important Biosynthetic Intermediates In aerobic organisms, the citric acid cycle is an amphibolic pathway, one that serves in both catabolic and anabolic processes. Role of the citric acid cycle in anabolism. 35 3. Gluconeogenesis The central role of glucose in metabolism--fuel and building block In mammals, some tissues depend almost completely on glucose for their metabolic energy (like brain and nervous system, erythrocytes, renal medulla). Supply of glucose from stores is not always sufficient, like during longer fasts, or after vigorous exercise. Through a pathway called gluconeogenesis, converts pyruvate and related three- and four-carbon compounds to glucose. Precursor 36 Gluconeogenesis and glycolysis are not identical pathways running in opposite directions, although they do share several steps. However, three reactions of glycolysis are essentially irreversible and cannot be used in gluconeogenesis: •conversion of pyruvate to phosphoenolpyruvate •conversion of fructose 1,6-bisphosphate to fructose 6-phosphate •conversion of glucose 6-phosphate to glucose 37 Conversion of Pyruvate to Phosphoenolpyruvate Requires Two Exergonic Reactions This reaction cannot occur by reversal of the pyruvate kinase reaction of glycolysis. Instead, the phosphorylation of pyruvate requires enzymes in both the cytosol and mitochondria. In mitochondria, pyruvate is converted to oxaloacetate in a biotin-requiring reaction catalyzed by pyruvate carboxylase. Mitochondrial membrane has no transporter for oxaloacetate In the cytosol, oxaloacetate is converted to phosphoenolpyruvate by PEP carboxykinase. 38 Gluconeogenesis Is Energetically Expensive, but Essential Citric Acid Cycle Intermediates and Many Amino Acids Are Glucogenic Citrate, isocitrate, -ketoglutarate, succinyl-CoA, succinate, fumarate, and malate—all are citric acid cycle intermediates that can undergo oxidation † to oxaloacetate. Some or all of the carbon atoms of most amino acids are ultimately catabolized to pyruvate or to intermediates of the citric acid cycle. In contrast, no net conversion of fatty acids to glucose occurs in mammals. 39 Glucogenic Amino Acids. †site of entry Chapter 6. Metabolism and its regulation Section II. Metabolic Regulation: Glucose and Glycogen BEIJING, METRO MAP METABOLIC PATHWAYS 40 1. The Metabolism of Glycogen in Animals In a wide range of organisms, excess glucose is converted to polymeric forms for storage—glycogen in vertebrates and many microorganisms, starch in plants. [may represent up to 10% of the weight of liver and 1% to 2% of the weight of muscle] The glycogen in muscle is there to provide a quick source of energy for either aerobic or anaerobic metabolism. Liver glycogen serves as a reservoir of glucose for other tissues when dietary glucose is not available Glycogen granules in a hepatocyte 41 (1) Glycogen Breakdown Is Catalyzed by Glycogen Phosphorylase Glucose units of glycogen enter glycolytic pathway through the action of three enzymes: glycogen phosphorylase, glycogen debranching enzyme, and phosphoglucomutase. Stop at the fourth glucose residues away from an (1 → 6) branch point (1→4) Transfer branches 42 Glucose 1-phosphate is converted to glucose 6-phosphate by phosphoglucomutase. The glucose 6-phosphate in skeletal muscle can enter glycolysis and serve as an energy source. In liver, it can be released into the blood when the blood glucose level drops, which need glucose 6-phosphatase with the active site on the lumenal side of the ER. 43 (2) The Sugar Nucleotide UDP-Glucose Donates Glucose for Glycogen Synthesis Many reactions in which hexoses are transformed or polymerized involve sugar nucleotides (active form). What’s the aim of this transformation? •Their formation is metabolically irreversible, contributing to the irreversible synthetic pathways in which they are intermediates. •Nucleotide moiety has many groups that can undergo noncovalent interactions with enzymes. •Nucleotidyl group is an excellent leaving group, facilitating nucleophilic attack by activating the sugar carbon to which it is attached. •By “tagging” some hexoses with nucleotidyl groups, cells can set them aside in a pool for glycogen synthesis. G 0’ = -19.2 kJ/mol 44 Glycogen synthesis takes place prominently in the liver and skeletal muscles. The starting point for synthesis of glycogen is glucose 6-phosphate. hexokinase I and II in muscle and hexokinase IV in liver phosphoglucomutase UDP-glucose pyrophosphorylase UDP-glucose is immediate donor of glucose residues in the reaction of glycogen synthase 45 Glycogen synthase cannot make the (1→6) bonds found at the branch points of glycogen; these are formed by the glycogen-branching enzyme, also called amylo (1→4) to (1→6) transglycosylase. The glycogen-branching enzyme catalyzes transfer of a terminal fragment of 6 or 7 glucose residues from the nonreducing end of a glycogen branch having at least 11 residues to the C-6 hydroxyl group of a glucose residue at a more interior position of the same or another glycogen chain, thus creating a new branch. 46 How is a new glycogen molecule initiated? A: Glycogenin primes the initial sugar residues in glycogen, and it also catalyze this biosynthesis reaction. Muscle glycogenin forms dimers. The substrate, UDPglucose (shown in red ball-and-stick), is bound to a Rossman fold near the amino terminus. Glycogenin catalyzes two distinct reactions. (1) Transfer of a glucose residue from UDP-glucose to the hydroxyl group of Tyr194 of glycogenin, catalyzed by the protein’s intrinsic glucosyltransferase activity; (2) The nascent chain is extended by the sequential addition of seven more glucose residues, each derived from UDP-glucose, catalyzed by the chain-extending activity of glycogenin. 47 At this point, glycogen synthase takes over, further extending the glycogen chain. Glycogenin remains buried within the particle, covalently attached to the single reducing end of the glycogen molecule. Structure of the glycogen particle. Starting at a central glycogenin molecule, glycogen chains (12 to 14 residues) extend in tiers. Inner chains have two (1→6) branches each. Chains in the outer tier are unbranched. There are 12 tiers in a mature glycogen particle (only 5 are shown here), consisting of about 55,000 glucose residues in a molecule of about 21 nm diameter and Mr 107. 48 2. Coordinated Regulation of Glycolysis and Gluconeogenesis Gluconeogenesis employs most of the enzymes that act in glycolysis, but it is not simply the reversal of glycolysis. Seven of the glycolytic reactions are freely reversible, other three reactions of glycolysis are so exergonic as to be irreversible: those catalyzed by hexokinase, PFK, and pyruvate kinase. 49 (1) Hexokinase Isozymes Are Affected Differently by Their Product There are four hexokinase isozymes (designated I to IV). I~III type in muscle, working for energy production, whereas IV in liver working for maintaining blood glucose homeostasis. At normal blood glucose concentration, hexokinase I~III acts at or near its maximal rate, whereas the activity of type IV is very low; When the blood glucose concentration is high, as it is after a meal rich in carbohydrates, excess glucose is transported into hepatocytes, where hexokinase IV converts it to glucose 6-phosphate. Km=0.1 mM glucose in blood 4~5 mM Km=10 mM 50 Hexokinase IV is subject to inhibition by the reversible binding of a regulatory protein specific to liver. Glucose causes dissociation of the regulatory protein from hexokinase IV, relieving the inhibition (immediately after meal). The binding is much tighter in the presence of the allosteric effector fructose 6-phosphate (blood glucose drops below 5 mM), draws this enzyme into nucleus. 51 (2) Phosphofructokinase-1 Is under Complex Allosteric Regulation In addition to its substrate-binding sites, PFK has several regulatory sites at which allosteric activators or inhibitors bind (e.g. ATP, citrate, fructose 2,6-bisphosphate). Substrate (fructose 1,6bisphosphate) Allosteric effector (ADP) 52 (3) Pyruvate Kinase Is Allosterically Inhibited by ATP Pyruvate kinase is allosterically inhibited by ATP, acetyl-CoA, long-chain fatty acids (signals for energy supply) and Ala (one-step transformation of pyruvate), and fructose 1,6-bisphosphate triggers its activation. In liver, glucagon inactivates pyruvate kinase (L isoenzyem) via phosphorylation. When the glucagon level drops, pyruvate kinase is activating via dephosphorylation. 53 (4) Gluconeogenesis Is Regulated at Several Steps The 1st control point, pyruvate can be converted to glucose and glycogen via gluconeogenesis or oxidized to acetyl-CoA for energy production. This enzyme in each path is regulated allosterically; acetyl-CoA, produced either by fatty acid oxidation or by the pyruvate dehydrogenase complex, stimulates pyruvate carboxylase and inhibits pyruvate dehydrogenase. 54 The second control point in gluconeogenesis is the reaction catalyzed by fructose 1,6bisphosphatase-1, which is strongly inhibited by AMP. The corresponding glycolytic enzyme, PFK-1, is stimulated by AMP and ADP but inhibited by citrate and ATP. In general, when sufficient concentrations of acetyl-CoA or citrate are present, or when a high energy charge, gluconeogenesis is favored. AMP promotes glycogen degradation and glycolysis by activating glycogen phosphorylase (via activation of phosphorylase kinase) and stimulating the activity of PFK-1. 55 3. Coordinated Regulation of Glycogen Synthesis and Breakdown (1) Regulation of Glycogen Phosphorylase Two interconvertible forms of glycogen phosphorylase, a (active) and b (inactive). Phosphorylase b predominates in resting muscle, but during vigorous muscular activity the epinephrine triggers phosphorylation of phosphorylase b, converting it to it’s a form, catalyzed by phosphorylase b kinase. Cascade mechanism of epinephrine and glucagon action. Glucose binding to an allosteric site of the phosphorylase a isozyme of liver induces a conformational change that exposes its phosphorylated Ser residues to the action of phosphorylase a phosphatase 1(PP1). phosphorylase a was converted to phosphorylase b. 56 (2) Glycogen Synthase Is Also Regulated by Phosphorylation and Dephosphorylation Two forms of glycogen Synthase: glycogen synthase a, active form, is unphosphorylated; glycogen synthase b, inactive, is phosphorylated (several sites). Glycogen synthase kinase 3 (GSK3) catalyzed the phosphorylation of glycogen synthase (three sites) and inactive it. 57 The action of GSK3 needs other protein kinase casein kinase II (CKII), has first phosphorylated the glycogen synthase on a nearby residue, an event called priming. In liver, conversion of glycogen synthase b to the active form is promoted by phosphorprotein phosphatase (PP1), which removes the phosphoryl groups. Glucose 6-phosphate binds to an allosteric site on glycogen synthase b, making the enzyme a better substrate for dephosphorylation by PP1 and causing its activation. 58 (3) Glycogen Synthase Kinase 3 Mediates the Actions of Insulin Insulin binding to its receptor activates a tyrosine protein kinase, which phosphorylates insulin receptor substrate-1 (IRS-1). IRS-1 is then bound by phosphatidylinositol 3-kinase (PI-3K), which converts phosphatidylinositol 4,5bisphosphate (PIP2) in to PIP3. tyrosine protein kinase A protein kinase (PDK-1) that is activated when bound to PIP3 activates a second protein kinase (PKB), which phosphorylates glycogen synthase kinase 3 (GSK3). The inactivation of GSK3 allows phosphoprotein phosphatase 1 (PP1) to dephosphorylate glycogen synthase (active form), therefore, stimulates glycogen synthesis 59 (4) Phosphoprotein Phosphatase 1 Is Central to Glycogen Metabolism •Phosphoprotein Phosphatase 1 can remove phosphoryl groups from phosphorylase kinase, glycogen phosphorylase, and glycogen synthase, which involved in response to glucagon and epinephrine. •PP1 does not exist free in the cytosol, but is tightly bound to glycogentargeting proteins (GM) that bind glycogen and above mentioned enzymes. Glycogen granule (1)Insulin-stimulated phosphorylation of GM site 1 activates PP1, which dephosphorylates phosphorylase kinase, glycogen phosphorylase, and glycogen synthase. (2) Epinephrine stimulated phosphorylation of GM site 2 causes dissociation of PP1 from the glycogen particle, preventing its access to glycogen phosphorylase and glycogen synthase. PKA also phosphorylates a protein (inhibitor 1) that, when phosphorylated, inhibits PP1. 60 (5) Allosteric and Hormonal Signals Coordinate Carbohydrate Metabolism Carbohydrate metabolism is well regulated by insulin, glucagon, and epinephrine. Elevated of blood glucose triggers insulin release. In hepatocyte, insulin has two immediate effects: it inactivates GSK3 and activates PP1, therefore, activates glycogen synthesis. High capacity PP1 inactivates glycogen transporter phosphorylase a and phosphorylase kinase, stopping glycogen breakdown. Glucose enters the hepatocyte through GLUT2, and the elevated intracellular glucose leads to hexokinase IV entering the cytosol and stimulating glycolysis and supplying the precursor for 61 glycogen synthesis. Low blood glucose triggers the release of glucagon, which activates PKA. This enzyme can •Phosphorylate phosphorylase kinase, activating it and leading to the activation of glycogen phosphorylase. •Phosphorylate glycogen synthase, inactivating it and blocking glycogen synthesis. •Phosphorylate PFK-2/FBPase-2, leading to a drop in the concentration of the regulator fructose 2,6-bisphosphate, which inactivate the glycolytic enzyme PFK-1 and activate the gluconeogenic enzyme FBPase-1. •Phosphorylate and inactivates the glycolytic enzyme pyruvate kinase. 62 Insulin Changes the Expression of Many Genes Involved in Carbohydrate and Fat Metabolism In addition to its effects on the activity of existing enzymes, insulin also regulates the expression of as many as 150 genes, including some related to fuel metabolism. 63 FURTHER READING: Simoni RD, Hill RL, Vaughan M. (2002) Carbohydrate metabolism: glycogen phosphorylase and the work of Carl F. and Gerty T. Cori. J. Biol. Chem. 277 (www.jbc.org/cgi/content/full/277/29/e18). Gibbons BJ, Roach PJ, Hurley TD. (2002) Crystal structure of the autocatalytic initiator of glycogen biosynthesis, glycogenin. J. Mol. Biol. 319, 463–477. Barford D. (1999) Structural studies of reversible protein phosphorylation and protein phosphatases. Biochem. Soc. Trans. 27, 751–766. de la Iglesia N, Mukhtar M, Seoane J, Guinovart JJ, Agius L. (2000) The role of the regulatory protein of glucokinase in the glucose sensory mechanism of the hepatocyte. J. Biol. Chem. 275, 10,597–10,603. Nordlie RC, Foster JD, Lange AJ. (1999) Regulation of glucose production by the liver. Annu. Rev. Nutr. 19, 379–406. Harwood AJ. (2001) Regulation of GSK-3: a cellular multiprocessor. Cell 105, 821–824. Radziuk J, Pye S. (2001) Hepatic glucose uptake, gluconeogenesis and the regulation of glycogen synthesis. Diabetes/Metab. Res. Rev. 17, 250–272. 64