After pyruvate is oxidized, the citric acid cycle completes the energy-yielding oxidation of organic molecules. Chapter 9, Section 3

Oxidation of Pyruvate to Acetyl CoA  In the presence of O 2, pyruvate enters the mitochondrion (in eukaryotic cells) where the oxidation of glucose is completed.  Before the citric acid cycle can begin, pyruvate must be converted to acetyl Coenzyme A (acetyl CoA), which links glycolysis to the citric acid cycle.  This step is carried out by a multienzyme complex that catalyses three reactions.

Figure 9.10 Pyruvate Transport protein CYTOSOL MITOCHONDRION CO 2 Coenzyme A NAD  + H  NADH Acetyl CoA 123

The Citric Acid Cycle  The citric acid cycle, also called the Krebs cycle, completes the break down of pyruvate to CO 2.  If acetyl-CoA is NOT stored then it enters the Citric Acid Cycle.  The cycle oxidizes organic fuel derived from pyruvate, generating 1 ATP, 3 NADH, and 1 FADH 2 per turn.

Figure Acetyl CoA Citrate Citric acid cycle CoA-SH Oxaloacetate Step One: A Condensation Reaction  2-C acetyl-CoA + 4-C molecule  6-C molecule+ CoA [which can be used over and over!].  Irreversible.  Inhibited by large amounts of ATP already present.

Figure Acetyl CoA Citrate Isocitrate Citric acid cycle H2OH2O 2 CoA-SH Oxaloacetate Step Two: Isomerization  Hydroxyl group repositioned.  Water removed from one carbon and then added to a different carbon.  RESULT: a change in position of an − H and − OH.  Molecule is still (6-C), but the –OH has moved!

Figure Acetyl CoA Citrate Isocitrate  -Ketoglutarate Citric acid cycle NADH + H  NAD  H2OH2O 32 CoA-SH CO2CO2 Oxaloacetate Step Three: First Oxidation  Molecule undergoes oxidative decarboxylation--fancy talk for chopping off a carbon and losing a pair of e - ’s in the process.  The pair of e - reduce NAD + to NADH.  The chopped off C becomes CO 2.  Now we have a 5-C molecule.

Figure Acetyl CoA Citrate Isocitrate  -Ketoglutarate Succinyl CoA Citric acid cycle NADH + H  NAD  H2OH2O 324 CoA-SH CO2CO2 CO2CO2 Oxaloacetate Step Four: Second Oxidation  The 5-C molecule undergoes oxidative decarboxylation.  Releases CO 2.  2 more e - reduce another NAD + to NADH  The 4-C fragment that is left receives a CoA group.

Figure Acetyl CoA Citrate Isocitrate  -Ketoglutarate Succinyl CoA Succinate Citric acid cycle NADH ATP + H  NAD  H2OH2O ADP GTPGDP P i 3245 CoA-SH CO2CO2 CO2CO2 Oxaloacetate Step Five: Substrate Level Phosphorylation  CoA leaves the 4-C molecule.  Breaking of bond releases energy.  GDP + P i  GTP [just substitute guanine for adenine in “A” TP].  GTP  ATP  Remaining 4-C molecule.

Figure Acetyl CoA Citrate Isocitrate  -Ketoglutarate Succinyl CoA Succinate Fumarate Citric acid cycle NADH FADH 2 ATP + H  NAD  H2OH2O ADP GTPGDP P i FAD CoA-SH CO2CO2 CO2CO2 Oxaloacetate Step Six: Third Oxidation  Four carbon molecule is oxidized.  FAD + + FOUR e - and TWO H +  FADH 2.  FAD + is an integral part of mitochondria membrane.  FADH 2 can contribute e - to ETS.

Figure NADH 1 Acetyl CoA Citrate Isocitrate  -Ketoglutarate Succinyl CoA Succinate Fumarate Malate Citric acid cycle NAD  NADH FADH 2 ATP + H  NAD  H2OH2O H2OH2O ADP GTPGDP P i FAD CoA-SH CO2CO2 CO2CO2 Oxaloacetate Step Seven and Eight: Regeneration  Water is added to oxidized 4-C molecule.  After water is added, compound becomes oxidized again (2 e - are released)  NAD + + the 2 e -  NADH  Ready to start again!

Figure 9.11 Pyruvate NAD  NADH + H  Acetyl CoA CO 2 CoA 2 CO 2 ADP + P i FADH 2 FAD ATP 3 NADH 3 NAD  Citric acid cycle + 3 H 

Beating a dead horse! Substrate Level Phosphorylation Oxidation Glycolysis 2 ATP 2 NADH Oxidation of Pyruvate NADH Krebs 2 ATP 6 NADH + 2 FADH 2 TOTAL4 ATP10 NADH + 2 FADH 2

Concept 9.4: During oxidative phosphorylation, chemiosmosis couples electron transport to ATP synthesis Following glycolysis and the citric acid cycle, NADH and FADH 2 account for most of the energy extracted from food. These two electron carriers donate electrons to the electron transport chain, which powers ATP synthesis via oxidative phosphorylation.

The Pathway of Electron Transport  The electron transport chain is in the inner membrane (cristae) of the mitochondrion.  Most of the chain’s components are proteins, which exist in multiprotein complexes.  The carriers alternate reduced and oxidized states as they accept and donate electrons.  Electrons drop in free energy as they go down the chain and are finally passed to O 2, forming H 2 O.

Figure 9.13 NADH FADH 2 2 H  + 1 / 2 O 2 2 e  H2OH2O NAD  Multiprotein complexes (originally from NADH or FADH 2 ) I II III IV Free energy (G) relative to O 2 (kcal/mol) FMN Fe  S FAD Q Cyt b Cyt c 1 Cyt c Cyt a Cyt a 3 Fe  S

 Electrons are transferred from NADH or FADH 2 to the electron transport chain.  Electrons are passed through a number of proteins including cytochromes (each with an iron atom) to oxygen.  The electron transport chain generates no ATP directly.  It breaks the large free-energy drop from food to O 2 into smaller steps that release energy in manageable amounts.

Chemiosmosis: The Energy-Coupling Mechanism  Electron transfer in the electron transport chain causes proteins to pump H + from the mitochondrial matrix to the intermembrane space.  H + then moves back across the membrane, passing through the protein, ATP synthase.  ATP synthase uses the exergonic flow of H + to drive phosphorylation of ADP.  This is an example of chemiosmosis, the use of energy in a H + gradient to drive cellular work.

Figure 9.14 INTERMEMBRANE SPACE Rotor Stator HH Internal rod Catalytic knob ADP + P i ATP MITOCHONDRIAL MATRIX

Figure 9.15 Protein complex of electron carriers (carrying electrons from food) Electron transport chain Oxidative phosphorylation Chemiosmosis ATP synth- ase I II III IV Q Cyt c FAD FADH 2 NADH ADP  P i NAD  HH 2 H  + 1 / 2 O 2 HH HH HH 21 HH H2OH2O ATP

 The energy stored in a H + gradient across a membrane couples the redox reactions of the electron transport chain to ATP synthesis.  The H + gradient is referred to as a proton-motive force, emphasizing its capacity to do work.

An Accounting of ATP Production by Cellular Respiration  During cellular respiration, most energy flows in this sequence: glucose  NADH  electron transport chain  proton-motive force  ATP  About 34% of the energy in a glucose molecule is transferred to ATP during cellular respiration, making about 32 ATP.  There are several reasons why the number of ATP is not known exactly.

Figure 9.16 Electron shuttles span membrane MITOCHONDRION 2 NADH 6 NADH 2 FADH 2 or  2 ATP  about 26 or 28 ATP Glycolysis Glucose 2 Pyruvate Pyruvate oxidation 2 Acetyl CoA Citric acid cycle Oxidative phosphorylation: electron transport and chemiosmosis CYTOSOL Maximum per glucose: About 30 or 32 ATP