Chapter 13 - The Citric Acid Cycle
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Transcript of Chapter 13 - The Citric Acid Cycle
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Chapter 13 - The Citric Acid CycleChapter 13 - The Citric Acid Cycle
• The citric acid cycle (tricarboxylic acid cycle) is amphibolic (both catabolic and anabolic)
• The cycle is involved in the aerobic catabolism of carbohydrates, lipids and amino acids
• Intermediates of the cycle are starting points for many biosynthetic reactions
• Enzymes of the cycle are in the mitochondria (eukaryotes) or the cytosol of bacteria
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Energy in the citric acid cycleEnergy in the citric acid cycle
• Energy of the oxidation reactions is largely conserved as reducing power
• Coenzymes reduced:
NAD+ NADH
Ubiquinone (Q) Ubiquinol (QH2)
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Entry of Pyruvate into the Entry of Pyruvate into the MitochondrionMitochondrion
• Pyruvate translocase transports pyruvate into the mitochondria in symport with H+
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Conversion of Pyruvate to Conversion of Pyruvate to Acetyl CoAAcetyl CoA
• Pyruvate dehydrogenase complex (PDH complex) is a multienzyme complex containing:
3 enzymes + 5 coenzymes + other proteins
(+ ATP coenzyme as a regulator)
E1 = pyruvate dehydrogenase
E2 = dihydrolipoamide acetyltransferase
E3 = dihydrolipoamide dehydrogenase
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Overall reaction of pyruvate Overall reaction of pyruvate dehydrogenase complexdehydrogenase complex
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The five steps of the PDH complexThe five steps of the PDH complexStep 1: Catalyzed by E1
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Step 2: The second step is also Step 2: The second step is also catalyzed by Ecatalyzed by E11
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Step 3: EStep 3: E22 transfers the lipoamide-bound acetyl transfers the lipoamide-bound acetyl
group to HS-CoA forming acetyl CoAgroup to HS-CoA forming acetyl CoA
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Step 4: EStep 4: E3 3 FAD group oxidizes reduced FAD group oxidizes reduced
lipoamide of Elipoamide of E22 forming FADH forming FADH22
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Step 5: EStep 5: E33-FADH-FADH22 reduces NAD reduces NAD++ to to
regenerate Eregenerate E33-FAD and NADH-FAD and NADH
• The oxidation of E3-FADH2 regenerates the original holoenzyme completing the catalytic cycle
• NADH dissociates from the complex
E3-FADH2 + NAD+ E3-FAD + NADH + H+
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Reactions of the PDH complexReactions of the PDH complex
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Roles of the coenzymes of the PDH Roles of the coenzymes of the PDH complexcomplex
• NAD+ and HS-CoA are cosubstrates
• TPP, lipoamide and FAD are prosthetic groups
• ATP is a regulator of the PDH complex
• Lipoamide (on E2) acts as a “swinging arm” to transfer the two carbon unit from the active site of E1 to the active site of E3 (substrate channeling)
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The Citric Acid Cycle The Citric Acid Cycle Oxidizes AcetylCoAOxidizes AcetylCoA
• Table 12.2
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Summary of the citric acid cycleSummary of the citric acid cycle
• For each acetyl CoA which enters the cycle:
(1) Two molecules of CO2 are released
(2) Coenzymes NAD+ and Q are reduced
(3) One GDP (or ADP) is phosphorylated
(4) The initial acceptor molecule (oxaloacetate) is reformed
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• Citric acid cycle
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• Fates of carbon atoms in the cycle
• Carbon atoms from acetyl CoA (red) are not lost in the first turn of the cycle
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Energy conservation by the cycleEnergy conservation by the cycle
• Energy is conserved in the reduced coenzymes NADH, QH2 and one GTP
• NADH, QH2 can be oxidized to produce ATP by oxidative phosphorylation
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The Citric Acid Cycle Can The Citric Acid Cycle Can Be a Multistep CatalystBe a Multistep Catalyst
• Oxaloacetate is regenerated
• The cycle is a mechanism for oxidizing acetyl CoA to CO2 by NAD+ and Q
• The cycle itself is not a pathway for a net degradation of any cycle intermediates
• Cycle intermediates can be shared with other pathways, which may lead to a resupply or net decrease in cycle intermediates
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1. Citrate Synthase1. Citrate Synthase• Citrate formed from acetyl CoA and oxaloacetate
• Only cycle reaction with C-C bond formation
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Proposed mechanism of citrate synthase Proposed mechanism of citrate synthase
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2. Aconitase2. Aconitase
• Elimination of H2O from citrate to form C=C bond of cis-aconitate
• Stereospecific addition of H2O to cis-aconitate to form 2R,3S-Isocitrate
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Reaction of AconitaseReaction of Aconitase
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Three point attachment of Three point attachment of prochiral substrates to enzymesprochiral substrates to enzymes
• Chemically identical groups a1 and a2 of a prochiral molecule can be distinguished by the enzyme
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• Fates of carbon atoms in the cycle
• Carbon atoms from acetyl CoA (red) are not lost in the first turn of the cycle
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3. Isocitrate Dehydrogenase3. Isocitrate Dehydrogenase
• Oxidative decarboxylation of isocitrate to-ketoglutarate (-kg) (a metabolically irreversible reaction)
• One of four oxidation-reduction reactions of the cycle
• Hydride ion from the C-2 of isocitrate is transferred to NAD+ to form NADH
• Oxalosuccinate is decarboxylated to -kg
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Isocitrate dehydrogenase reactionIsocitrate dehydrogenase reaction
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4. The a-Ketoglutarate Dehydrogenase 4. The a-Ketoglutarate Dehydrogenase ComplexComplex
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Structure of a-Ketoglutarate Structure of a-Ketoglutarate dehydrogenase complexdehydrogenase complex
• Similar to pyruvate dehydrogenase complex
• Same coenzymes, identical mechanisms
E1 - -ketoglutarate dehydrogenase (with TPP)
E2 - succinyltransferase (with flexible lipoamide prosthetic group)
E3 - dihydrolipoamide dehydrogenase (with FAD)
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5. Succinyl-CoA Synthetase5. Succinyl-CoA Synthetase• Free energy in thioester bond of succinyl CoA
is conserved as GTP (or ATP in plants, some bacteria)
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• Mechanism of succinyl-CoA synthetase (continued on next slide)
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6. The Succinate 6. The Succinate Dehydrogenase (SDH) ComplexDehydrogenase (SDH) Complex
• Located on the inner mitochondrial membrane (other components are dissolved in the matrix)
• Dehydrogenation is stereospecific; only the trans isomer is formed
• Substrate analog malonate is a competitive inhibitor of the SDH complex
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Reaction of the succinate Reaction of the succinate dehydrogenase complexdehydrogenase complex
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Succinate and malonateSuccinate and malonate
• Malonate is a structural analog of succinate
• Malonate binds to the enzyme active site, and is a competitive inhibitor
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Structure of the SDH complexStructure of the SDH complex
• Complex of several polypeptides, an FAD prosthetic group and iron-sulfur clusters
• Electrons are transferred from succinate to ubiquinone (Q), a lipid-soluble mobile carrier of reducing power
• FADH2 generated is reoxidized by Q
• QH2 is released as a mobile product
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7. Fumarase7. Fumarase
• Stereospecific trans addition of water to the double bond of fumarate to form L-malate
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8. Malate Dehydrogenase8. Malate Dehydrogenase
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Reduced Coenzymes Fuel the Reduced Coenzymes Fuel the Production of ATPProduction of ATP
• Each acetyl CoA entering the cycle nets:
(1) 3 NADH
(2) 1 QH2
(3) 1 GTP (or 1 ATP)
• Oxidation of each NADH yields 2.5 ATP
• Oxidation of each QH2 yields 1.5 ATP
• Complete oxidation of 1 acetyl CoA = 10 ATP
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Glucose degradation via glycolysis, citric Glucose degradation via glycolysis, citric acid cycle, and oxidative phosphorylationacid cycle, and oxidative phosphorylation
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Regulation of the Citric Acid CycleRegulation of the Citric Acid Cycle
• Pathway controlled by:
(1) Allosteric modulators
(2) Covalent modification of cycle enzymes
(3) Supply of acetyl CoA
(4) Regulation of pyruvate dehydrogenase complex controls acetyl CoA supply
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Regulation of the PDH complexRegulation of the PDH complex
• Increased levels of acetyl CoA and NADH inhibit E2, E3 in mammals and E. coli
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Regulation of mammalian PDH Regulation of mammalian PDH complex by complex by covalentcovalent modificationmodification
• Phosphorylation/dephosphorylation of E1
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Further regulation of the PDH complexFurther regulation of the PDH complex
Pyruvate dehydrogenase kinase (PDK)
• PDK is activated by NADH and acetyl CoA (leads to inactivation of the PDH complex)
• PDK is inhibited by pyruvate and ADP (leads to activation of the PDH complex)
Pyruvate dehydrogenase phosphatase (PDP)
• PDP activity is stimulated by Ca2+ (leads to an activation of the PDH complex)
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Control points in the citric acid cycle
Rate is adjusted to meet the cell’s need for ATP. Three allosteric enzyme control points:
PDH - inhibited by NADH, acetyl CoA, and ATP.
Isocitrate dehydrogenase - stimulated by ADP; inhibited by ATP and NADH
a-ketoglutarate dehydrogenase—inhibited by NADH, succinyl CoA, high energy charge.
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The Glyoxylate CycleThe Glyoxylate Cycle• Pathway for the formation of glucose from
noncarbohydrate precursors in plants, bacteria and yeast (not animals)
• Glyoxylate cycle leads from 2-carbon compounds to glucose
• In animals, acetyl CoA is not a carbon source for the net formation of glucose (2 carbons of acetyl CoA enter cycle, 2 are released as 2 CO2)
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Glyoxylate cycle - formation of glucoseGlyoxylate cycle - formation of glucose
• Formation of glucose from acetyl CoA (or any substrate that is a precursor to acetyl CoA)
• Ethanol or acetate can be metabolized to acetyl CoA and then to glucose via the glyoxylate cycle
• Stored seed oils in plants are converted to carbohydrates during germination
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Isocitrate lyase: first bypass Isocitrate lyase: first bypass enzyme of glyoxylateenzyme of glyoxylate
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Malate synthase: second bypass Malate synthase: second bypass enzyme of glyoxylateenzyme of glyoxylate
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Glyoxylate cycle in germinating Glyoxylate cycle in germinating castor beanscastor beans
• Conversion of acetyl CoA to glucose requires the transfer of metabolites among three metabolic compartments
(1) The glyoxysome(2) The cytosol(3) The mitochondrion