Gluconeogenesis UNIT II: Intermediary Metabolism.
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- Gluconeogenesis UNIT II: Intermediary Metabolism
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- Figure 10.1. The gluconeogenesis pathway shown as part of the essential pathways of energy metabolism. The numbered reactions are unique to gluconeogenesis..
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- Overview Some tissues e.g., brain, RBCs, kidney medulla, lens & cornea, testes, & exercising muscle, require a continuous supply of gluc as a metabolic fuel. Liver glycogen, an essential postprandial source of gluc, can meet these needs for only 10-18 h in the absence of dietary intake of CHO During a prolonged fast, hepatic glycogen stores are depleted, & gluc is formed from precursors such as lactate, pyruvate, glycerol (from backbone of triglycerols), and -ketoacids (from catabolism of glucogenic aas) The formation of gluc does not occur by simple reversal of glycolysis, because overall equil of glycolysis strongly favors pyruvate formation Instead gluc is synthesized by a special pathway, gluconeogenesis During an o/n fast, ~ 90% of gluconeogenesis occurs in liver, with kidneys providing 10% of the newly synthesized gluc molecules However, during prolonged fasting, kidneys become major gluc- producing organs, contributing an estimated 40% of the total gluc production
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- II. Substrates for gluconeogenesis -Gluconeogenic precursors are molecules that can be used to produce a net synthesis of gluc. -They include all the intermediates of glycolysis and TCA cycle. Glycerol, lactate, and -keto acids obtained from deamination of glucogenic aas are the most important gluconeogenic precursors A. Glycerol -Is released during the hydrolysis of triglycerols in adipose tissue, & is delivered by blood to the liver -Glycerol is phosphorylated by glycerol kinase to glycerol-P, which is oxidized by glycerol phosphate dehydrogenase to DHAP = an intermediate of glycolysis Note: adipocytes cant phosphorylate glycerol because they lack glycerol kinase
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- B. Lactate -Lactate is released into blood by exercising skeletal muscle, and by cells that lack mitoch e.g., RBCs -In the Cori cycle, blood- borne gluc is converted by exercising muscle to lactate, which diffuses into the blood. This lactate is taken up by the liver and converted to gluc, which is released back into circulation Figure 10.2. The Cori cycle.
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- C. Amino acids -Amino acids derived from hydrolysis of tissue proteins are the major sources of gluc during a fast. --ketoacids, e.g., OAA and -KG, are derived from the metabolism of glucogenic amino acids. -These substances can enter the TCA cycle and form OAA, a direct precursor of PEP Note: Acetyl CoA & cpds that give rise to acetyl CoA (e.g., acetoacetate and aas such as Lys and Leu) cant give rise to a net synthesis of gluc. This is due to the irreversible nature of the pyruvate dehydrogenase reaction, which converts pyruvate to acetyl CoA. These cpds give rise to ketone bodies and are therefore temred ketogenic
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- III. Reactions unique to gluconeogenesis Seven glycolytic reactions are reversible and are used in the synthesis of gluc from pyruvate or lactate. However, 3 of the reactions are irreversible and must be circumvented by 4 alternate reactions that energetically favor synthesis of gluc A. Carboxylation of pyruvate -The 1 st roadblock to overcome in synthesis of gluc from pyruvate is the irreversible conversion in glycolysis of pyruvate to PEP by pyruvate kinase -In gluconeogenesis pyruvate is 1 st carboxylated by pyruvate carboxylase to OAA, which is then converted to PEP by PEP-carboxykinase
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- Figure 10.3. Activation and transfer of CO2 to pyruvate, followed by transport of oxaloacetate to the cytosol and subsequent decarboxylation.
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- 1. Biotin is a coenzyme: -Pyruvate carboxylase contains biotin, which is covalently bound to the enz protein through the - amino group of Lys, forming an active enz -This covalently bound form of biotin is called biocytin -Cleavage of a high-energy phosphate of ATP drives formation of an enz-biotin-CO2 intermediate. This high-energy complex subsequently carboxylates pyruvate to form OAA Note: this reaction occurs in the mitoch of liver & kidney cells, & has 2 purposes: to provide an important substrate for gluconeogenesis, & to provide OAA that can replenish TCA cycle intermediates that may become depleted, depending on synthetic needs of the cell. Muscle cells also contain pyruvate carboxylase, but use OAA produced only for the latter purpose, they do not synthesize glucose.
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- 2. Allosteric regulation: - Pyruvate carboxylase is allosterically activated by acetyl CoA. Elevated levels of acetyl CoA may signal one of several metabolic states in which the increased synthesis of OAA is required. - e.g., this may occur during fasting in which OAA is used for synthesis of gluc by gluconeogenesis in the liver & kidney. - Conversely, at low levels of acetyl CoA, pyruvate carboxylase is largely inactive, & pyruvate is primarily oxidized by pyruvate dehydrogenase to produce acetyl CoA that can be further oxidized by the TCA cycle.
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- B. Transport of OAA to the cytosol -OAA, formed in mitoch., must enter cytosol where the other enzs of gluconeogenesis are located. -However, OAA is unable to directly cross inner mitoch memb; it must first be reduced to malate by mitoch malate dehydrogenase -Malate can be transported from mitoch to cytosol, where it is reoxidized to OAA by cytosol malate dehydrogenase C. Decarboxylation of cytoslic OAA -OAA is decarboxylated & phosphorylated in cytosol by PEP-carboxykinase (= PEPCK). -The reaction is driven by hydrolysis of GTP. -The combined actions of pyruvate carboxylase & PEPCK provide an energetically favorable pathway from pyruvate to PEP. -PEP is then acted on by the reactions of glycolysis running in the reverse direction until it becomes F-1,6-BP
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- D. Dephosphorylation of fructose 1,6-bisphosphate -Hydrolysis of F-1,6-BP by fructose 1,6-bisphosphatase bypasses the irreversible PFK-1 reaction, & provides an energetically favorable pathway for formation of F- 6-P. This reaction is an important regulatory site of gluconeogenesis 1. Regulation by energy levels within the cell: - fructose 1,6-bisphosphatase is inhibited by elevated levels of AMP, which signal an energy-poor state in the cell. Conversely, high levels of ATP & low concs of AMP stimulate gluconeogenesis 2. Regulation by fructose 2,6-bisphosphate: - Fructose 1,6-bisphosphatase, found in liver & kidney, is inhibited by fructose 2,6-bisphosphate, an allosteric modifier whose conc is influenced by the level of circulating glucagon Note: recall that F-2,6-BP activates PFK-1 of glycolysis, thus allowing for reciprocal control of gluc synthesis & oxidation
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- Figure 10.4 Dephosphorylation of fructose 1,6- bisphosphate.
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- Figure 10.5. Effect of elevated glucagon on the intracellular concentration of fructose 2,6-bisphosphate in the liver. PFK-2 = phosphofructokinase-2; FBP-2 = Fructose bisphospate phosphatase-2.
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- E. Dephosphorylation of glucose 6-phosphate -Hydrolysis of G-6-P by glucose 6-phosphatase bypasses the irreversible hexokinase reaction, & provides an energetically favorable pathway for the formation of free gluc. -Liver & kidney are the only organs that release free gluc from G-6-P -This process actually requires 2 enzs: glucos 6- phosphate translocase, which transports G-6-P across the ER memb, & a 2 nd ER enz, glucose 6-phosphatase (found only in gluconeogenic cells), which removes P, producing free gluc.
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- Figure 10.6. Dephosphorylation of glucose 6-phosphate.
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- Note: these enzs are required for the final steps in glycogenolysis, as well as gluconeogenesis - Type Ia glycogen storage disease (Von Gierke disease) results from an inherited deficiency of one of these enzymes - Specific transporters are responsible for releasing free gluc & P back into cytosol, & in hepatocytes, into the blood Note: muscle lacks glucose 6-phosphatase & therefore, cant provide blood gluc by gluconeogenesis. Also, G-6- P derived from muscle glycogen cant be dephosphorylated to yield free gluc.
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- F. Summary of the reactions of glycolysis & gluconeogenesis -Of the 11 reactions required to convert pyruvate to free gluc, 7 are catalyzed by reversible glycolytic enzs. -The irreversible reactions of glycolysis catalyzed by hexokinase, PFK, & pyruvate kinase are circumvented by glucose 6-phosphatase, fructose 1,6-bisphosphatase & pyruvate carboxylase/PEP carboxykinase -In gluconeogenesis, the equilibria of the 7 reactions of glycolysis are pushed in favor of gluc synthesis as a result of the essentially irreversible formation of PEP, F-6-P, & glucose catalyzed by gluconeogenic enzs. Note: stoichiometry of gluconeogenesis from pyruvate couples cleavage of 6 high-energy phosphate bonds & oxidation of 2 NADH with formation of each molecule of glucose
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- Figure 10.7. Summary of the reactions of glycolysis and gluconeogenesis, showing the energy requirements of gluconeogenesis.
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- IV. Regulation of gluconeogenesis The moment-to-moment regulation of gluconeogenesis is determined primarily by the circ