Lecture 8 (2009)

Lecture 8 (2009) - Lecture 8 Tricarboxylic Acid (TCA) Cycle...

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Unformatted text preview: Lecture 8 Tricarboxylic Acid (TCA) Cycle (Citric Acid Cycle; Krebs Cycle) Oxidation of Acetyl-CoA to CO2 Generation of NADH and FADH2 Substrate-level phosphorylation Overall Goal: C6H12O6 + 6O2 6CO2 + 6H2O CARBOHYDRATES Glucose Amino acids Fatty acids Other Carbohydrates Glycolysis Glycogen Glucose-6-P Pyruvate PDH TCA Cycle O2 Acetyl-CoA NADH ATP CO2 (2x) Lactate CO2 (4x) H2O (1 x Glucose) p. 21 Glucose 2NAD+ 2H2O 2ATP 2 Pyruvate 2NADH + H+ Cytosol Glycolysis 2 Pyruvate 2NAD+ 2CoA-SH Mitochondria PDH Complex 2CO2 2 Acetyl~CoA 2NADH + H+ The PDH Complex (revisited) localized in mitochondrial matrix (5 co-factors required) largest multienzyme complex known (larger than the ribosome) 60 E1 subunits; 60 E2 subunits; 12 E3 subunits subunits subunits Advantage: substrate channeling suppression of side reactions concerted regulation Acetyl-CoA ("activated acetate") Thioester (high free energy of hydrolysis) Acidic protons S C H O P Reactive carbon atom N Acetyl- Coenzyme A Mitochondria 2 Acetyl~CoA 6NAD+ 2FAD 4H2O 2ATP 2FADH2 6NADH + H+ TCA Cycle (8 reactions) 4 CO2 Why are so many reactions required? Why are they arranged in a cycle? Mitochondria 2 Acetyl~CoA 6NAD+ 2FAD 4H2O 2ATP 2FADH2 6NADH + H+ TCA Cycle (8 reactions) 4 CO2 1. Logistics of C-C bond cleavage (not a simple feat) 2. Requirement of sequential oxidation steps 3. Dual function (catabolic and anabolic, or amphibolic) H H C C CoA-S H How to break a C-C bond? O In biological systems, C-C cleavage is assisted by a proximal carbonyl group (in "" position) or analogous group. C C C TCA cycle intermediates provide a "scaffold" O The Tricarboxylic Acid (TCA) Cycle CoA-SH + H+ ACETYL-CoA H2O O C ScoA H3C CH2COOHO COOCH2COOCITRATE H2O CH2COOC COOCHCOO - H2O 2 2 1 CH2COOH COOHC COOOH NAD ISOCITRATE cis-aconitate enzyme-bound O C COOH2C COOOXALOACETATE NADH + H+ 8 NAD 3 NADH + H CO2 CH2COOH H C COOO -KETOGLUTARATE COO HO C H H C H COOMALATE 7 CoASH 5 - 4 NAD + CoA-SH NADH + H + CO2 H -OOC C C COOH FADH2 6 H2O COOCH2 CH2 COOFAD SUCCINATE CH2COOH H C ScoA O GTP GDP + Pi SUCCINYL-CoA FUMARATE p. 45 Reaction 1: Citrate synthase (aldol condensation; hydrolysis of thioester) Go' = - 32 kJ mol-1 CoA-SH + H+ ACETYL-CoA H2O O H3C C ScoA The importance of "activated" acetate (Acetyl-CoA) CH2COOHO COOCH2COOCITRATE 1 O C COO H2C COO- OXALOACETATE p. 45 Reaction 2: Aconitase (Isomerization) Go' = + 13 kJ mol-1 + CH2 HO COO2 H2O COOCH2COOCITRATE CH2COO C COOCHCOO- - H2O 2 CH2COOH COOHC COOOH ISOCITRATE cis-aconitate enzyme-bound Tertiary "alcohol" Secondary "alcohol" p. 45 Reaction 3: Isocitrate Dehydrogenase (IDH) Go' = - 21 kJ mol-1 CH2COOH COOHC COOOH NAD 3 ISOCITRATE Oxidative Decarboxylation NADH + H CO2 CH2COOH H C COOO -KETOGLUTARATE p. 45 Reaction 4: -Ketoglutarate Dehydrogenase (KDH) CH2COOH H C COOO Go' = - 33 kJ mol-1 -KETOGLUTARATE 4 NAD + CoA-SH NADH + H + CO2 Oxidative Decarboxylation (similar to PDH complex) CH2COOH H C ScoA O SUCCINYL-CoA p. 45 Reaction 5: Succinyl-CoA Synthetase (Substrate-level phosphorylation) Go' = - 3 kJ mol-1 COO CH2 CH2 COOGTP SUCCINATE - CoASH 5 CH2COOH H C ScoA O GDP + Pi SUCCINYL-CoA GTP + ADP GDP + ATP Nucleoside diphosphate kinase (Go' = 0 kJ mol-1) p. 45 Reaction 6: Succinate Dehydrogenase (SDH) Go' = 0 kJ mol-1 COOHO C H H C H COOMALATE 7 Oxidoreduction H -OOC COOH FADH2 FUMARATE FAD SUCCINATE 6 C C H2O COOCH2 CH2 COO- p. 45 Reaction 7: Fumarase Go' = - 4 kJ mol-1 COOHO C H H C H COOMALATE Addition of H2O (stereospecific, only L-Malate) 7 H -OOC C C COOH FADH2 6 H2O COOCH2 CH2 COOFAD SUCCINATE FUMARATE p. 45 Reaction 8: Malate Dehydrogenase (MDH) ACETYL-CoA CoA-SH + H+ H2O CH2COOHO COOCH2COOCITRATE O H3C C ScoA 1 O C COOH2C COOOXALOACETATE NADH + H+ 8 Go' = + 30 kJ mol-1 NAD COOHO C H H C H COOMALATE p. 45 Reaction 1: Citrate synthase ACETYL-CoA CoA-SH + H+ H2O CH2COOHO COOCH2COOCITRATE Go' = - 32 kJ mol-1 O H3C C ScoA 1 O C COOH2C COOOXALOACETATE NADH + H+ Reaction 8: MDH Go' = + 30 kJ mol-1 NAD 8 COOHO C H H C H COOMALATE p. 45 The Tricarboxylic Acid (TCA) Cycle CoA-SH + H+ ACETYL-CoA H2O O H3C C ScoA CH2COOHO COOCH2COOCITRATE H2O CH2COOC COOCHCOO - H2O 2 2 1 CH2COOH COOHC COOOH NAD ISOCITRATE cis-aconitate enzyme-bound O C COOH2C COOOXALOACETATE NADH + H+ 8 NAD 3 NADH + H CO2 CH2COOH H C COOO -KETOGLUTARATE COO HO C H H C H COOMALATE 7 CoASH 5 - 4 NAD + CoA-SH NADH + H + CO2 H -OOC C C COOH FADH2 6 H2O COOCH2 CH2 COOFAD SUCCINATE CH2COOH H C ScoA O GTP GDP + Pi SUCCINYL-CoA FUMARATE p. 45 Round 1 Acetyl-CoA Isocitrate OAA CO2 Succinate CO2 -KGA Round 2 Acetyl-CoA Isocitrate OAA CO2 Succinate CO2 -KGA Round 3 Acetyl-CoA Isocitrate OAA CO2 Succinate CO2 -KGA Round 1 Acetyl-CoA Isocitrate OAA CO2 Succinate CO2 -KGA Total ATP Glucose 2 Pyruvate 2 ATP 2 NADH 2+6 2 Pyruvate 2 Acetyl-CoA + 2 CO2 4 CO2 2 ATP 2 NADH 6 2 Acetyl-CoA 6 NADH 2 FADH2 2 + 18 4 "Oxidative" ADP phosphorylation: 1 NADH = 3 ATP 1 FADH2 = 2 ATP 38 ATP The Pasteur Effect Glucose Glycolysis Glucose-6-P Pyruvate PDH TCA Cycle Acetyl-CoA NADH ETC O2 ATP CO2 CO2 H2O 38 ATP Aerobic Conditions: Low Glucose Consumption Glucose Glucose Glucose Glucose Glucose Glucose The Pasteur Effect Glycolysis Glucose-6-P Pyruvate 2 ATP Ethanol Anaerobic Conditions: High Glucose Consumption The Tricarboxylic Acid (TCA) Cycle CoA-SH + H+ ACETYL-CoA H2O O H3C C ScoA CH2COOHO COOCH2COOCITRATE H2O CH2COOC COOCHCOO - H2O 2 2 1 CH2COOH COOHC COOOH NAD ISOCITRATE cis-aconitate enzyme-bound O C COOH2C COOOXALOACETATE NADH + H+ 8 NAD 3 NADH + H CO2 CH2COOH H C COOO -KETOGLUTARATE COO HO C H H C H COOMALATE 7 CoASH 5 - 4 NAD + CoA-SH NADH + H + CO2 H -OOC C C COOH FADH2 6 H2O COOCH2 CH2 COOFAD SUCCINATE CH2COOH H C ScoA O GTP GDP + Pi SUCCINYL-CoA FUMARATE p. 45 Common "Metabolic Motifs" of the TCA Cycle -KGA Reactions 1-3 OAA Introduction of (-CH2-) group "Chain elongation" (e.g., amino acid biosynthesis) Decarboxylation of -ketoacids (e.g., pyruvate) Substrate-level phosphorylation (ATP) ATP Reaction 4 -KGA-DH Reaction 5 Succinyl-CoA synthetase Reactions 6-8 Succinate OAA Introduction of a keto function (e.g., fatty acid degradation) Fuel Energy Action Food Energy Regeneration Reproduction Carbon Chemistry Biosynthetic Functions of the TCA Cycle Steroids PEP Pyruvate Fatty Acids Acetyl-CoA Pyrimidine Bases Acetyl-CoA (cytosol) Thr Asp Oxalaoacetate Citrate Ile Met Lys Asn Malate TCA Cycle Isocitrate Glutathione -Ketoglutarate Fumarate Glu Ornithine Succinyl-CoA Pro Gln Arg Porphyrines Purine Bases Heme Chlorophyll Vitamin B12 p. 46 Anabolic Function Glycolysis PEP Pyruvate Catabolic Function CO2 Acetyl-CoA Oxalaoacetate Citrate Malate TCA Cycle Isocitrate CO2 Fumarate -Ketoglutarate Remove Intermediates Succinyl-CoA CO2 Anabolic Function Glycolysis PEP Pyruvate Catabolic Function CO2 Acetyl-CoA Oxalaoacetate Citrate Malate TCA Cycle Isocitrate CO2 Fumarate -Ketoglutarate Remove Intermediates Succinyl-CoA CO2 Anabolic Function Glycolysis PEP Pyruvate Catabolic Function CO2 Anaplerotic Reactions CO2 C3 Acetyl-CoA C2 Oxalaoacetate Citrate Malate TCA Cycle Isocitrate CO2 Fumarate -Ketoglutarate Remove Intermediates Succinyl-CoA CO2 Anaplerotic Reactions 14 CO2 H2O H214CO3 Carbonic acid H+ H14CO3Bicarbonate -O -O C O H 14 CO3- ATP ADP + HPO4 2- C C O O O CH3 Pyruvate Carboxylase (biotin-dependent, only in mitochondria of animals) CH2 14COO- Pyruvate Oxaloacetate p. 47 Biotin O C HN HC H2C S NH CH C H (CH2)4 O C H N (CH2)4 NH CH C O Lysine residue of enzyme (isopeptide bond) p. 48 O C O O C N HC H2C S NH CH C H Rest N-Carboxybiotin p. 48 Anaplerotic Reactions 14 CO2 H2O H214CO3 Carbonic acid H+ H14CO3Bicarbonate -O -O C O H 14 CO3- ATP ADP + HPO4 2- C C O O O CH3 Pyruvate Carboxylase (biotin-dependent, only in mitochondria of animals) CH2 14COO- Pyruvate Oxaloacetate p. 47 -O H14CO3C O HPO42- -O C C O O OPO32CH2 PEP Carboxylase (bacteria, yeast, plants [cyt.], not animals) CH2 14 COO- Phosphoenolpyruvate (PEP) Oxaloacetate (OAA) -O - C C CH2 14 O GTP GDP + 14 CO2 O C O O OPO32- COO- PEP Carboxykinase (PEPCK) (localization varies among species: only mitochondrial, only cytosolic, or both) CH2 OAA PEP p. 47 ...
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