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Download Biochem 2 Recitation #2 Spr 20152102105.pptx
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Biochem 2 Recitation #2 Glycolysis & Gluconeogenesis How do transporters exert control on metabolic pathways? There are several fates for glucose when it enters a cell; • These fates are all following the initial rxn that forms glucose 6-P04. • The pathways that claim G6P are; pentose phosphate pathway, glycogen synthesis pathway as well as entering glycolysis. • The final product of glycolysis is pyruvate. In mammalian cells it has two major fates, homolactate fermentation or aerobic respiration. Glycolysis can be thought of as consisting of three separate portions. • Stage 1 is the phosphorylation of glucose and the isomerization to Fr-6-P04; why is this step necessary? • Stage 2 is the isomerization of GAP and DHAP. Why is DHAP the favored product? • Stage 3 is the utilization of GAP to form pyruvate. What are the controls of stage 3? Glucose + 2NAD à 2pyruvate + NADH ΔG0’ = -146 kJ/mol 2ADP + 2Pià 2ATP + H2O ΔG0’ = 61 kJ/mol Overall free energy -85 kJ/mol Glycolysis is not a reversible pathway. Gluconeogensis then must be a separate pathway. Glucoseà2 Lactate ΔG0= -196 kJ/mol But the lactate dh is reversible, there are 5 isoenzymes of Ldh in most mammals. Why are gluconeogensis and glycolysis reciprocal pathways? • Three major enzymes are not reversible due to their thermodymanics, these are; • Hexokinase • PFK-1 • Pyruvate kinase. • Gluconeogenesis is a reciprocal pathway to glycolysis, and not a mere reversal of glycolysis. At specific highly exothermic reaction in glycolysis, the reverse reaction is not possible. A second enzyme is necessary that reverses that reaction step. There are three steps of this kind, Pyruvate Kinase, PFK-1/PFK-2, and Hexokinase/Glucokinase. • Non- carbohydrate precursors of glucose via gluconeogenesis are lactate, pyruvate, amino acids (primarily alanine) and Krebs cycle intermediates (especially OAA/malate, and αKG). Regulation of hexokinase IV (glucokinase) by sequestration in the nucleus. The protein inhibitor of hexokinase IV is a nuclear binding protein that draws hexokinase IV into the nucleus when the fructose 6-phosphate concentration in liver is high and releases it to the cytosol when the glucose concentration is high. • This is the committed step to glycolysis, the product F16BP is the only molecule that can be cleaved by the reverse Claisen condensation rxn, catalzyed by aldolase A to form GAP + DHAP. • PFK-1 activity is controlled by the product of PFK-2, the homotropic activator F26BP. Fr2,6BP is not a glycolytic substrate, and found in µM conc. The glycerol-3-PO4 Shuttle • the enzyme called cytoplasmic glycerol-3-phosphate dehydrogenase (cGPdh) converts DHAP to glycerol 3-phosphate by oxidizing one molecule of NADH to NAD+ • Glycerol-3-phosphate gets converted back to DHAP by a membrane-bound mGPdh, this time reducing one molecule of enzyme-bound FAD to FADH2. FADH2 then reduces coenzyme Q (ubiquinone to ubiquinol) which enters into oxidative phosphorylation. This reaction is irreversible Regulation of PK. The enzyme is allosterically inhibited by ATP, acetyl-CoA, and long-chain fa (all signs of an abundant energy supply), and the accumulation of fr 1,6-BP triggers its activation. Accumulation of alanine, which can be synthesized from pyruvate in one step, allosterically inhibits PK, slowing the production of pyruvate by glycolysis. The liver isozyme (L form) is also regulated hormonally. Glucagon activates cAMP-dependent protein kinase (PKA), which phosphorylates the PK L isozyme, inactivating it. When the glucagon level drops, a protein phosphatase (PP) dephosphorylates PK, activating it. This mechanism prevents the liver from consuming glucose by glycolysis when blood glucose is low; instead, the liver exports glucose. The muscle isozyme (M form) is not affected by this phosphorylation mechanism. Transport of PEP and oxaloacetate from the mitochondrion to the cytosol. The participation of two second messenger systems: the cAMP-mediated stimulation of glycogenolysis and inhibition of glycogen synthesis triggered by glucagon and beta-andrenoreceptor activation; and the IP3, DAG, and Ca2+-mediated stimulation of glycogenolysis and inhibition of glycogen synthesis triggered by alpha-adrenoreceptor activation. IP3 stimulates the release of Ca2+ from the endoplasmic, whereas DAG, together with Ca2+, activates protein kinase C(PKC) to phosphorylate and thereby inactivate glycogen synthase. G6Pase occupies the endoplasmic reticulum. Consequently, the cytosolically produced G6P is transported into the endoplasmic reticulum via the T1 G6P translocase, where it is hydrolyzed to glucose and Pi. The glucose and Pi are then returned to the cytosol by the T2 and T3 transporters, respectively, and the glucose is exported from the cell via the GLUT2 glucose transporter.