LECTURE PRESENTATIONS For CAMPBELL BIOLOGY NINTH EDITION Jane
LECTURE PRESENTATIONS For CAMPBELL BIOLOGY, NINTH EDITION Jane B. Reece, Lisa A. Urry, Michael L. Cain, Steven A. Wasserman, Peter V. Minorsky, Robert B. Jackson Chapter 9 Cellular Respiration and Fermentation Lectures by Erin Barley Kathleen Fitzpatrick © 2011 Pearson Education, Inc.
Overview: Life Is Work • living cells require energy from outside sources • some animals obtain energy by eating plants • some animals feed on other organisms that eat plants • energy flows into an ecosystem as sunlight and leaves as heat • photosynthesis generates O 2 and organic molecules - used in cellular respiration • cells use chemical energy stored in organic molecules to regenerate ATP - powers work Light energy ECOSYSTEM CO 2 H 2 O Photosynthesis in chloroplasts Cellular respiration in mitochondria Organic O 2 molecules ATP powers most cellular work Heat energy
Catabolic Pathways and Production of ATP • catabolism - the breakdown of organic molecules is exergonic – potential energy is stored in chemical bonds – result of the arrangement of electrons in these bonds – breaking of these bonds releases this potential energy • catabolic processes – anaerobic, fermentation & aerobic • Fermentation = partial degradation of sugars that occurs without O 2 • Aerobic respiration consumes organic molecules (fuel) and O 2 to yield ATP – most prevalent and efficient catabolic process – carried out by eukaryotic cells and most prokaryotic • Anaerobic respiration - similar to aerobic respiration but consumes compounds other than O 2 – prokaryotic cells
• Cellular respiration includes both aerobic and anaerobic respiration – glycolysis, Kreb’s cycle and electron transport chain – is often used to refer to aerobic respiration • uses carbohydrates, fats, and proteins • most cellular respiration starts with the sugar glucose C 6 H 12 O 6 + 6 O 2 6 CO 2 + 6 H 2 O + Energy (ATP + heat) • breakdown of glucose is exergonic – ΔG = -686 kcal/mol • creates ATP – powers the work of endergonic processes
Redox Reactions: Oxidation and Reduction • How do catabolic pathways decompose glucose and yield energy • through the transfer of electrons = oxidationreduction reactions or Redox reactions – relocation of electrons releases stored potential energy • this released energy is ultimately used to synthesize ATP
The Principle of Redox • redox reactions = chemical reactions that transfer electrons between reactants • oxidation = a substance loses electrons, or is oxidized • reduction = a substance gains electrons, or is reduced (the amount of positive charge is reduced) becomes oxidized becomes reduced becomes oxidized (loses electron) becomes reduced (gains electron) • the electron donor is called the reducing agent – e. g. Xe- = reduces Y • the electron receptor is called the oxidizing agent – e. g. Y = oxidizes Xe by removing its electrons
Reactants becomes oxidized Products Energy becomes reduced Methane (reducing agent) Oxygen (oxidizing agent) Carbon dioxide Water • the problem becomes when you have covalent bonds • i. e. not all redox reactions involve the complete transfer of electrons from one substance to another – but change the degree of electron sharing in covalent bonds • an example is the reaction between methane and O 2 – in methane – covalent electrons are shared equally between the C and the Hs of methane and the two Os of O 2 - they are equally electronegative – BUT - O is a very electronegative element – one of the most potent oxidizing agents – when CO 2 is formed - electrons are shared less equally between C and O – the O “pulls” on the electrons harder – so the carbon has lost electrons (becomes oxidized) – the electrons are also shared less equally in the water – O has gained electrons & become reduced
Reactants becomes oxidized Products Energy becomes reduced Methane (reducing agent) Oxygen (oxidizing agent) Carbon dioxide Water • energy must be added to pull an electron away from an atom • the more electronegative an atom is – the more energy will be required for it to become oxidized – like rolling a ball uphill – methane is not very electronegative – little energy is required for it to become oxidized • an electron loses potential energy as it shifts from a less electronegative atom to a more electronegative atom – just as a ball loses potential energy when it rolls downhill – it releases this loss of potential energy as chemical energy - used for work
• an electron loses potential energy as it shifts from a less electronegative atom to a more electronegative atom • a redox reaction that moves electrons closer to oxygen – releases chemical energy – e. g. form CO 2 from methane – electrons move from the Carbon (when in methane) closer to the Oxygen of CO 2 = ENERGY
Oxidation of Organic Fuel Molecules During Cellular Respiration • During cellular respiration, the fuel (such as glucose) is oxidized, and O 2 is reduced becomes oxidized becomes reduced • the carbon atoms of glucose are not very electronegative • lose their potential energy when they combine with O 2 to form CO 2 – move away from C and closer to O (more electronegative) • in general – molecules with carbons are good sources of potential energy – electrons will shift toward more electronegative atoms like oxygen
Oxidation of Organic Fuel Molecules During Cellular Respiration becomes oxidized becomes reduced • in respiration – the oxidation of glucose (loss of electrons) transfers electrons to a lower energy state (lower G) – as electrons move from a less electronegative compound (sugar) to a more electronegative compound (CO 2) – this liberates energy – used to make ATP • only the activation energy barrier prevents the transfer of electrons to the lower energy state • body temperature is not high enough to reach this activation energy barrier – done by enzymes • main sources of ATP in cells – carbohydrates and fats
Stepwise Energy Harvest via NAD+ and the Electron Transport Chain • so oxidizing glucose produces more electronegative compounds and releases energy • if you release this energy all at once – not very efficient • in cellular respiration - glucose and other organic molecules are broken down to these electronegative compounds in a series of steps – at key steps – electrons are stripped from glucose and eventually transferred to O 2 – in typical oxidation reactions - electrons that are transferred travel with a proton (H+)
NAD+ • BUT electrons are not transferred directly – first transferred to NAD+ - functions as a coenzyme • NAD = nicotinamine adenine dinucleotide • as an electron acceptor- NAD+ functions as an oxidizing agent during cellular respiration • each NADH (the reduced form of NAD+) represents stored energy that is tapped to synthesize ATP
NAD NADH Dehydrogenase Reduction of NAD (from food) Nicotinamide (oxidized form) Oxidation of NADH Nicotinamide (reduced form) • How does NAD+ trap electrons? • transfer is dependent upon a kind of enzyme called a Dehydrogenase • dehydrogenases remove a pair of hydrogen atoms and a pair of electrons • ACTUALLY – dehydrogenase removes a hydride ion/H- (two electrons and a proton) and a proton (H+)
Figure 9. UN 04 NADH Dehydrogenase Reduction of NAD (from food) Oxidation of NADH Nicotinamide (oxidized form) Nicotinamide (reduced form) • one electron gets transferred onto the N+ of NAD N • the second electron gets attached to the C 4 carbon directly across from the N • this is REDUCTION and produces a reduced form of NAD (NAD-) • one proton attaches to the nicotinamide ring NADH • the other is released into the surroundings Dehydrogenase
• the electrons lose very little of their potential energy when they are transferred to NAD+ • each NADH represents stored energy that can be tapped to make ATP when the electrons “fall” down an energy gradient to oxygen 2 H 1/ 2 O 2 (from food via NADH) Free energy, G ATP rt spo Explosive release of heat and light energy ATP tran tron ain ch Free energy, G 2 H+ 2 e Controlled release of energy for synthesis of ATP Elec • cellular respiration uses an Electron Transport Chain to break this fall into several energy releasing steps • NADH shuttles the electrons to the top of the hill • O 2 captures these electrons along with H+ to form water H 2 1/2 O 2 2 e 2 H+ H 2 O (a) Uncontrolled reaction H 2 O (b) Cellular respiration
• electron transfer from NADH to O 2 = -53 kcal/mol • use of several redox reactions to allow electrons to fall from one carrier to another 2 H 1/ 2 O 2 (from food via NADH) Free energy, G ATP rt spo Explosive release of heat and light energy ATP tran tron ain ch Free energy, G 2 H+ 2 e Controlled release of energy for synthesis of ATP Elec • each “downhill” carrier is more electronegative than its “uphill” carrier & capable of oxidizing it • electrons eventually fall down the ETC to a far more stable location in an oxygen atom H 2 1/2 O 2 2 e 2 H+ H 2 O (a) Uncontrolled reaction H 2 O (b) Cellular respiration
The Stages of Cellular Respiration • Harvesting of energy from glucose has three stages: – Glycolysis –anaerobic process that breaks down glucose into two molecules of pyruvate • cytosol – the citric acid cycle (Kreb’s Cycle) - completes the breakdown of glucose • starting material = acetyl Co. A (derived from pyruvate) • matrix of the mitochondria (cytosol in prokaryotes) – Oxidative phosphorylation = ETC and chemiosmosis • generates most of the ATP • powered by redox reactions • inner mitochondrial membrane (plasma membrane in prokaryotes) Electrons carried via NADH and FADH 2 Electrons carried via NADH Glycolysis Glucose Pyruvate CYTOSOL Pyruvate oxidation Acetyl Co. A Citric acid cycle Oxidative phosphorylation: electron transport and chemiosmosis MITOCHONDRION ATP ATP Substrate-level phosphorylation Oxidative phosphorylation
• oxidative phosphorylation accounts for almost 90% of the ATP generated by cellular respiration • a smaller amount of ATP is formed in glycolysis and the citric acid cycle by substrate-level phosphorylation • ALSO = some of the steps of Glycolysis and Citric Acid Cycle are redox reactions involving dehydrogenases removing protons and electrons and creating NADH • for each molecule of glucose degraded to CO 2 and water by respiration- the cell makes up to 36 molecules of ATP – 4 by substrate level phosphorylation Enzyme ADP P Substrate ATP Product
Glycolysis harvests chemical energy by oxidizing glucose to pyruvate Energy Investment Phase • Glycolysis (“splitting of sugar”) breaks down glucose into two molecules of pyruvate • Glycolysis has two major phases – Energy investment phase – Energy payoff phase • Glycolysis occurs whether or not O 2 is present Glucose 2 ADP 2 P 2 ATP used Energy Payoff Phase 4 ADP 4 P 2 NAD+ 4 e 4 H+ 4 ATP formed 2 NADH 2 H+ 2 Pyruvate 2 H 2 O Net Glucose 4 ATP formed 2 ATP used 2 NAD+ 4 e 4 H+ 2 Pyruvate 2 H 2 O 2 ATP 2 NADH 2 H+
Figure 9. 9 -1 Glycolysis: Energy Investment Phase Glucose ATP Glucose 6 -phosphate ADP Hexokinase 1 1. Hexokinase – transfers a phosphate group from ATP to glucose-6 -phosphate - makes the glucose it more chemically reactive
Figure 9. 9 -2 Glycolysis: Energy Investment Phase Glucose ATP Glucose 6 -phosphate Fructose 6 -phosphate ADP Hexokinase 1 Phosphoglucoisomerase 2 2. Phosphoglucoisomerase – enzyme that catalyzes the rearrangement of G 6 P into fructose-6 -phosphate
Figure 9. 9 -3 Glycolysis: Energy Investment Phase Glucose ATP Glucose 6 -phosphate Fructose 6 -phosphate ATP Fructose 1, 6 -bisphosphate ADP Hexokinase 1 Phosphoglucoisomerase Phosphofructokinase 2 3 3. Phosphofructokinase – transfers a phosphate group from ATP to the opposite end of F 6 P fructose-1, 6 -bisphosphate -phosphofructokinase is the rate-limiting enzyme for the glycolytic portion of cellular respiration
Figure 9. 9 -3 Glycolysis: Energy Investment Phase Glucose ATP Glucose 6 -phosphate Fructose 6 -phosphate ATP Fructose 1, 6 -bisphosphate ADP Hexokinase 1 Phosphoglucoisomerase Phosphofructokinase 2 3 -phosphofructokinase catalyzes an irreversible step in glycolysis -same is true for hexokinase and pyruvate kinase -its activity is regulated by the reversible binding of an allosteric effector -inhibited by high levels of ATP – acts to lower its affinity for fructose-6 -phosphate -ATP binds to the enzyme at a regulatory site – changes the shape of the enzyme’s catalytic site -reduced binding to F 6 P results -enzyme can also can be inhibited through a drop in p. H
Figure 9. 9 -4 Glycolysis: Energy Investment Phase Glucose ATP Glucose 6 -phosphate Fructose 6 -phosphate ATP ADP Hexokinase 1 Fructose 1, 6 -bisphosphate Phosphoglucoisomerase Phosphofructokinase 2 3 Aldolase Dihydroxyacetone phosphate 4 Glyceraldehyde 3 -phosphate Isomerase 5 4. Aldolase – cleaves the 6 carbon F-1, 6 -BP into two 3 -Carbon sugars dihydroxyacetone phosphate and glyceraldehyde-3 phosphate - are isomers of each other -DHAP is a ketose, G 3 P is an aldose -if this could reach equilibrium – 96% would be DHAP -BUT G 3 P is used as soon as it is formed – never reaches Eq. To step 6
Figure 9. 9 -4 Glycolysis: Energy Investment Phase Glucose ATP Glucose 6 -phosphate Fructose 6 -phosphate ATP ADP Hexokinase 1 Fructose 1, 6 -bisphosphate Phosphoglucoisomerase Phosphofructokinase 2 3 Aldolase Dihydroxyacetone phosphate 4 Glyceraldehyde 3 -phosphate Isomerase 5 To step 6 5. Triose phosphate Isomerase – converts DAHP G 3 P immediately after G 3 P is used -G 3 P forms the main glycolytic pathway so the isomerase funnels DHAP into this pathway by converting it into G 3 P -the following reactions actually run twice!!! (two molecules of G 3 P are produced)
Figure 9. 9 -5 Glycolysis: Energy Payoff Phase 2 NADH 2 NAD Triose phosphate dehydrogenase 6 + 2 H 2 Pi 6. Triose phosphate Dehydrogenase -also known as Glyceraldehyde-3 -phosphate dehydrogenase (GAPDH) -first step: removes 2 electrons and 2 protons 1, 3 -Bisphoglycerate from G 3 P and transfers them to NAD+ to make NADH -so G 3 P is oxidized and NAD+ is reduced -transfer of electrons creates a more electronegative compound (NAD+) releases energy -second step: energy released by this redox reaction is used for phosphorylation of G 3 P 1, 3 -bisphoglycerate
Figure 9. 9 -6 Glycolysis: Energy Payoff Phase 2 ATP 2 NADH 2 NAD Triose phosphate dehydrogenase 6 + 2 H 2 ADP 2 Phosphoglycerokinase 2 Pi 1, 3 -Bisphoglycerate 7 3 -Phosphoglycerate 7. Phosphoglycerokinase: – 1, 3 -BPG is unstable and has a high energy potential -allows for the transfer of the phosphate group from C 1 to ADP ATP is made via Substrate level Phosphorylation -1, 3 -BPG is oxidized 3 -phosphoglycerate (when the phosphate group is bound to 1, 3 -BPG - one electron is pulled closer to the O of the sugar – O is more electronegative than P – when the P group is released, it takes it’s electron & the sugar loses it – i. e. oxidized)
Figure 9. 9 -7 Glycolysis: Energy Payoff Phase 2 ATP 2 NADH 2 NAD Triose phosphate dehydrogenase 6 + 2 H 2 ADP 2 2 Phosphoglyceromutase Phosphoglycerokinase 2 Pi 1, 3 -Bisphoglycerate 7 3 -Phosphoglycerate 8 2 -Phosphoglycerate 8. Phosphoglyceromutase: transfer of the remaining phosphate group from C 3 to C 2 2 -phosphoglycerate
Figure 9. 9 -8 Glycolysis: Energy Payoff Phase 2 ATP 2 NADH 2 NAD Triose phosphate dehydrogenase 6 + 2 H 2 ADP 2 H 2 O 2 2 1, 3 -Bisphoglycerate 7 Enolase Phosphoglyceromutase Phosphoglycerokinase 2 Pi 2 3 -Phosphoglycerate 8 2 -Phosphoglycerate 9 Phosphoenolpyruvate (PEP) 9. Enolase: extracts a water molecule from the sugar and forms a double bond between C 2 and C 3 phosphoenolpyruvate (PEP)
Figure 9. 9 -9 Glycolysis: Energy Payoff Phase 2 ATP 2 NADH 2 NAD Triose phosphate dehydrogenase 6 + 2 H 2 ADP 2 H 2 O 2 2 1, 3 -Bisphoglycerate 7 3 -Phosphoglycerate 8 2 ADP 2 Enolase Phosphoglyceromutase Phosphoglycerokinase 2 Pi 2 ATP 2 -Phosphoglycerate 9 Pyruvate kinase Phosphoenolpyruvate (PEP) 10 Pyruvate 10. Pyrvuate kinase: PEP is unstable and has a high energy potential -allows the transfer of the remaining P group to ADP for the second round of Substrate Level Phosphorylation ATP is made -this final step of glycolysis produces Pyruvate
Glycolysis Energy Tally • 2 NADH from glycolysis = 4 ATP from ETC • 4 ATP made through substrate phosphorylation • 2 ATP consumed
After pyruvate is oxidized, the citric acid cycle completes the energy-yielding oxidation of organic molecules • glycolysis releases less than a quarter of the chemical energy in glucose • most energy is still contained in pyruvate • oxidation of pyruvate releases the remaining energy • in eukaryotes and in the presence of O 2 - pyruvate enters the mitochondrion - the oxidation of glucose is completed • takes place in the cytosol in prokaryotes
Transition Phase: Oxidation of Pyruvate to Acetyl Co. A • before the citric acid cycle can begin- pyruvate must be converted to acetyl Coenzyme A (acetyl Co. A) – links glycolysis to the citric acid cycle • this transition step is carried out by a multienzyme complex in the matrix that catalyzes three reactions = pyruvate dehydrogenase complex • requires a transport pump to bring pyruvate into the matrix of the mitochondria – powered by the H+ gradient of the electron transport chain 1. removal of the carboxyl CYTOSOL group as a molecule of CO 2 2 carbon compound 2. oxidation into acetate & transfer of electrons to NAD+ 3. coenzyme A attached via its sulfur atom acetyl Pyruvate co. A Transport protein MITOCHONDRION CO 2 Coenzyme A 3 1 2 NADH + H Acetyl Co. A
Transition Phase Energy Tally • 2 NADH from pyruvate oxidation = 6 ATP from ETC
Fermentation and anaerobic respiration enable cells to produce ATP without the use of oxygen • in the absence of oxygen, glycolysis couples with fermentation or anaerobic respiration to produce ATP • Fermentation uses substrate-level phosphorylation instead of an electron transport chain to generate ATP – Fermentation consists of glycolysis plus reactions that regenerate NAD +, which can be reused by glycolysis • Anaerobic respiration uses an electron transport chain with a final electron acceptor other than O 2, for example sulfate
Fermentation • Two common types are alcohol fermentation and lactic acid fermentation • in alcohol fermentation - pyruvate is converted to ethanol in two steps – – first step generates CO 2 second step: acetylaldehyde ethanol + NAD+ catalyzed by Alcohol Dehydrogenase Alcohol fermentation by yeast is used in brewing, winemaking, and baking 2 ADP 2 P i Glucose 2 ATP Glycolysis 2 Pyruvate 2 NAD 2 Ethanol (a) Alcohol fermentation 2 NADH 2 CO 2 2 Acetaldehyde
• in lactic acid fermentation - pyruvate is reduced by NADH, forming lactate as an end product, with no release of CO 2 • Lactic acid fermentation by some fungi and bacteria is used to make cheese and yogurt • Human muscle cells use lactic acid fermentation to generate ATP when O 2 is scarce 2 ADP 2 P i Glucose 2 ATP Glycolysis 2 NAD 2 Lactate (b) Lactic acid fermentation 2 NADH 2 Pyruvate
“Fates” of Pyruvate • 1. ethanol • 2. lactate • 3. acetyl co. A • the production of lactate or ethanol is a reduction reaction and regenerates NAD+ • this sustains glycolysis by replenishing NAD+
• 4. PEP glucose • 5. amino acids • 6. oxaloacetate citric acid cycle
Fermentation and anaerobic respiration enable cells to produce ATP without the use of oxygen • Most cellular respiration requires O 2 to produce ATP • Without O 2, the electron transport chain will cease to operate • In that case, glycolysis couples with fermentation or anaerobic respiration to produce ATP • Anaerobic respiration uses an electron transport chain with a final electron acceptor other than O 2, for example sulfate • Fermentation uses substrate-level phosphorylation instead of an electron transport chain to generate ATP
• Obligate anaerobes carry out fermentation or anaerobic respiration and cannot survive in the presence of O 2 • Yeast and many bacteria are facultative anaerobes, meaning that they can survive using either fermentation or cellular respiration • In a facultative anaerobe, pyruvate is a fork in the metabolic road that leads to two alternative catabolic routes Glucose Glycolysis CYTOSOL Pyruvate No O 2 present: Fermentation Ethanol, lactate, or other products O 2 present: Aerobic cellular respiration MITOCHONDRION Acetyl Co. A Citric acid cycle
The Citric Acid Cycle Pyruvate • citric acid cycle- also called the Krebs cycle • completes the break down of pyruvate to CO 2 NAD Co. A NADH + H Acetyl Co. A – two more CO 2 produced per pyruvate – 6 total (including 2 from transition) • generates 1 GTP/ATP, 3 NADH, and 1 FADH 2 per turn • turns once for every pyruvate made • 2 pyruvates are made for each glucose Co. A Citric acid cycle 2 CO 2 3 NAD FADH 2 3 NADH FAD + 3 H ADP + P i ATP
The Citric Acid Cycle • The citric acid cycle has eight steps, each catalyzed by a specific enzyme • two carbons (red) enter in the reduced form as an acetyl group • two different carbons (blue) leave as the oxidized form of CO 2 • The acetyl group of acetyl Co. A joins the cycle by combining with oxaloacetate, forming citrate • The next seven steps decompose the citrate back to oxaloacetate, making the process a cycle • The NADH and FADH 2 produced by the cycle relay electrons extracted from food to the electron transport chain
Acetyl Co. A-SH 1 Oxaloacetate Citric acid cycle 1. Addition of Acetyl co. A to oxaloacetate to produce citrate -catalyzed by Citrate synthase or Co. A Synthase
Acetyl Co. A-SH H 2 O 1 Oxaloacetate 2 Citrate Isocitrate Citric acid cycle 2. Isomerization of citrate to iso-citrate: dehydration reaction, followed by a hydration step creates this isomer of citrate -intermediate step creates cis-Aconitate (unstable)
Figure 9. 12 -3 Acetyl Co. A-SH H 2 O 1 Oxaloacetate 2 Citrate Isocitrate NAD Citric acid cycle 3 NADH + H CO 2 -Ketoglutarate 3. Oxidation of isocitrate alpha-ketoglutarate: : 1 st of the 4 redox reactions in the citric acid cycle -creates an unstable intermediate that loses a CO 2 (oxidation) -coupled to the reduction of NAD+ to NADH - catalyzed by isocitrate dehydrogenase
Acetyl Co. A-SH H 2 O 1 Oxaloacetate 4. Oxidation of -ketoglutarate succinyl co. A: 2 nd of the 4 redox reactions in the citric acid cycle -loss of another CO 2 in the oxidation reaction -reduction of NAD+ to NADH -Co. A is added through an unstable bond – ΔG is -8 kcal/mol -catalyzed by -ketoglutarate dehydrogenase -3 enzyme complex -reactions similar to the creation of acetyl co. A 2 Citrate Isocitrate NAD Citric acid cycle NADH 3 + H CO 2 Co. A-SH -Ketoglutarate 4 NADH Succinyl Co. A + H CO 2
Acetyl Co. A-SH 5. Conversion to succinate: the energy released by the cleavage of the Co. A powers the phosphorylation of GDP GTP -the GTP can be used to generate another ATP through phosphate transfer -catalyzed by Succinyl Co. A synthase (similar enzyme as that added Co. A to oxaloacetate) H 2 O 1 Oxaloacetate 2 Citrate Isocitrate NAD Citric acid cycle NADH 3 + H CO 2 Co. A-SH -Ketoglutarate 4 Co. A-SH 5 NAD Succinate GTP GDP CO 2 NADH Pi Succinyl Co. A + H ADP ATP -in other cells – ATP is made at this step -bacteria, plants & some animal cells
Acetyl Co. A-SH 6 8: Oxidation of succinate to oxaloacetate: regeneration of oxaloacetate is done in three steps 6. Oxidation to Fumarate – reduction of a FADH 2 -FAD is used because the free energy change is not enough to reduce NAD+ 7. Hydration to Malate – addition of water rearranges fumarate into malate 8. Oxidation to oxaloacetate – coupled to reduction of another NAD+ NADH H 2 O 1 + H NAD 8 Oxaloacetate 2 Malate Citrate Isocitrate NAD H 2 O Citric acid cycle 7 Fumarate NADH 3 + H CO 2 Co. A-SH -Ketoglutarate 6 4 Co. A-SH 5 FADH 2 NAD FAD Succinate GTP GDP ATP Pi Succinyl Co. A NADH + H CO 2
Acetyl Co. A-SH NADH CITRIC ACID CYCLE TALLY -for each acetyl co. A entering the cycle H 2 O 1 + H NAD 8 Oxaloacetate 2 Malate 1. 3 NAD+ reduced to NADH (6 total = 18 ATP) 2. 1 FAD reduced to FADH 2 (2 total = 4 ATP) 3. production of 1 GTP (2 total = 2 ATP in some cells) 4. generation of 2 CO 2 (4 total) Citrate Isocitrate NAD H 2 O Citric acid cycle 7 Fumarate NADH 3 + H CO 2 Co. A-SH -Ketoglutarate 6 4 Co. A-SH 5 FADH 2 NAD FAD Succinate GTP GDP ATP Pi Succinyl Co. A NADH + H CO 2
Cellular Respiration = Theoretical Energy • Glucose contains 686 kcal/mol (up to 720 kcal/mol) • ATP contains 7. 3 kcal/mol • 36 ATP harvested from each molecule of glucose • 2 from glycolysis – substrate level phosphorylation • 2 from Kreb’s cycle – converted from GTP (not done by all cells) • 32 from the ETC & NADH/FADH 2 • 2 NADH – glycolysis • 8 NADH – transition + Kreb’s • 2 FADH 2 – Kreb’s • note: NADH in the cytosol is only worth 2 ATP; NADH in the mitochondria are worth 3 ATP • 36 moles of ATP x 7. 3 kcal/mol = 263 kcal trapped as ATP • efficiency : 263/686 = 38% efficiency in converting the potential energy of glucose into chemical energy
Glycolysis and the citric acid cycle connect to many other metabolic pathways Proteins Carbohydrates Amino acids Sugars Glycolysis Glucose Glyceraldehyde 3 - P NH 3 Pyruvate Acetyl Co. A Citric acid cycle Oxidative phosphorylation Fats Glycerol Fatty acids
Biosynthesis (Anabolic Pathways) • the body uses small molecules to build other substances • these small molecules may come directly from food, from glycolysis, or from the citric acid cycle • about 10 of the 20 amino acids can be made by converting substrates from the citric acid cycle or from pyruvate – rest are called essential amino acids • glucose can be made from pyruvate – gluconeogenesis • fatty acids can be made from acetyl co. A
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 their electrons to the electron transport chain - powers ATP synthesis via oxidative phosphorylation • donation of electrons to O 2 powers the phosphorylation of ADP ATP • enzyme of the electron transport chain is in the inner mitochodrial membrane (cristae) • most of the chain’s components are multiprotein complexes associated with carriers – 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
– the release free energy is broken up into smaller steps through the development of separate protein complexes • the “fall” is spontaneous as the uphill complex has a higher G value vs. the downhill • as the electrons fall they reduce the next downhill complex • eventually O 2 is reduced 50 2 e NAD FADH 2 2 e Free energy (G) relative to O 2 (kcal/mol) • the electron transport chain generates no ATP directly • ATP is generated as a result in the drop of free energy as the electrons “fall” from one protein complex to the next NADH 40 FMN I Fe S II Q III Cyt b 30 Multiprotein complexes FAD Fe S Cyt c 1 IV Cyt c Cyt a 20 10 0 Cyt a 3 2 e (originally from NADH or FADH 2) 2 H + 1/2 O 2 H 2 O
50 2 e NAD FADH 2 2 e Free energy (G) relative to O 2 (kcal/mol) • the ETC couples electron transfer to the generation of a proton-motive force for ATP synthesis NADH 40 FMN I Fe S II Q III Cyt b 30 Multiprotein complexes FAD Fe S Cyt c 1 IV Cyt c Cyt a 20 10 0 Cyt a 3 2 e (originally from NADH or FADH 2) 2 H + 1/2 O 2 H 2 O
– flavoprotein • Complex II – Succinate dehydrogenase – adds additional electrons to the ETC from succinate via FADH 2 • Complex III – Cytochrome bc 1 – also know as cytochrome c reductase • Complex IV – Cytochrome c oxidase • complexes I, II and III have ironsulfur groups (heme groups) that are critical to their function NADH 50 2 e NAD FADH 2 2 e Free energy (G) relative to O 2 (kcal/mol) • four protein complexes coupled to specific prosthetic groups: • Complex I – NADH dehydrogenase 40 FMN I Fe S II Q III Cyt b 30 Multiprotein complexes FAD Fe S Cyt c 1 IV Cyt c Cyt a 20 10 0 Cyt a 3 2 e (originally from NADH or FADH 2) 2 H + 1/2 O 2 H 2 O
NADH 50 2 e NAD FADH 2 2 e Free energy (G) relative to O 2 (kcal/mol) • Complex I – NADH dehydrogenase – flavoprotein because one of its prosthetic groups is flavin mononucleotide (FMN) – 4 H+ pumped into intermembrane space – 1. 2 electrons transferred to FMNH 2 (reduction in one step) – 2. Oxidation of FMNH 2 (in 2 steps) QH 2 (ubiquinol) • the two electrons moved via a Fe. S group Q (ubiquinone) • step A: 1 st electron converts Q semiquinone • step B: 2 nd electron converts semiquinone into ubiquinol (QH 2) • QH 2 is lipid soluble and diffuses through the membrane to Complex III 40 FMN I Fe S II Q III Cyt b 30 Multiprotein complexes FAD Fe S Cyt c 1 IV Cyt c Cyt a 3 20 2 e 10 (originally from NADH or FADH 2) 2 H + 1/2 O 2 0 QH 2 carries 2 electrons H 2 O
• Complex III – Cytochrome bc 1 – transfers its electrons via Q-Cycle • 4 H+ pumped as a result – QH 2 binds to the Qo site on the complex – Q binds to a second site called Qi – two electrons from QH 2 take two pathways: – path 1. one electron move to Fe-S group and then to a c 1 site • electron then reduces the cytochrome c 1 carrier and then a cytochrome c – path 2. 2 nd electron moves to a cytochrome b. L site then to a b. H site –> the Qi site of the complex • this electron moves from Qi site to reduce Q to semiquinone • it will take another cycle for this semiquinone to be reduced back to QH 2 • this QH 2 can then re-enter the Q cycle • so 2 cytochrome c’s are reduced and transferred to Complex IV • 4 protons are pumped into the intermembrane space This complex can “leak” electrons to O 2 and create superoxide radicals (water-soluble & found in the intermembrane space) THIS PROTEIN COMPLEX NEEDS 2 MOLECULES OF QH 2 TO RUN (4 electrons) -moves 4 H+
NADH 50 2 e NAD FADH 2 2 e Free energy (G) relative to O 2 (kcal/mol) • Complex IV – Cytochrome c Oxidase – 4 molecules of cyto c deliver their electrons to this complex – 4 electrons are transferred to cyto a group cyto a 3 group O 2 to create two molecules of water – 2 H+ are pumped by this complex – water production: – 2 e- + 2 H+ + ½ O 2 H 20 – 4 e- + 4 H+ + O 2 2 H 20 – this complex can be inhibited by cyanide 40 FMN I Fe S II Q III Cyt b 30 Multiprotein complexes FAD Fe S Cyt c 1 IV Cyt c Cyt a 20 10 0 Cyt a 3 2 e (originally from NADH or FADH 2) 2 H + 1/2 O 2 H 2 O
NADH 50 2 e NAD FADH 2 2 e Free energy (G) relative to O 2 (kcal/mol) • Complex II –Succinate Dehydrogenase – transfers the electrons produced through the reduction of FADH 2 – 2 electrons moved to Q to create QH 2 40 FMN I Fe S II Q III Cyt b 30 Multiprotein complexes FAD Fe S Cyt c 1 IV Cyt c Cyt a 20 10 0 Cyt a 3 2 e (originally from NADH or FADH 2) 2 H + 1/2 O 2 H 2 O
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 proton, ATP synthase • ATP synthase uses the exergonic flow of H+ to drive phosphorylation of ATP • This is an example of chemiosmosis, the use of energy in a H+ gradient to drive cellular work
• 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 H H H Protein complex of electron carriers Q I IV III II FADH 2 NADH H Cyt c FAD 2 H + 1/2 O 2 NAD ATP synthase H 2 O ADP P i (carrying electrons from food) ATP H 1 Electron transport chain Oxidative phosphorylation 2 Chemiosmosis
ATP-synthase – comprised of a: 1. stator – two “½ channels” for H+ binding 2. rotor – multiple H+ binding sites 3. internal rod 4. catalytic knob INTERMEMBRANE SPACE H Stator Rotor 1. H+ ion flowing down its gradient enters the 1 st half-channel in the stator 2. H+ ions enter binding sites within the rotor -binding of the H+ ions changes the rotors Internal rod shape so it rotates 3. each H+ ion makes one complete turn Catalytic knob before leaving the rotor and passing through the second half-channel within the stator 4. spinning of the rotor, rotates the rod and then the ADP knob below it + 5. rotation of the knob activates catalytic sites that Pi add a P group to ADP ATP MITOCHONDRIAL MATRIX http: //www. youtube. com/watch? v=3 y 1 d. O 4 n. Na. KY
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
An Accounting of ATP Production by Cellular Respiration • There are several reasons why the number of ATP - not known exactly – 1. the ratio of NADH molecules to ATP is not a whole number – phosphorylation and redox are not directly coupled to each other – 2. NADH made in the cytosol cannot enter the mitochondria • must use a “shuttle” to take these electrons into the matrix • number of ATP made depends on whether FAD or NAD+ is used as that shuttle – 3. use of the proton-motive force can be used for other mitochondrial work • powers the uptake of pyruvate from the cytosol • if this motive force was used exclusively for oxidative phosphorylation = 30 to 32 ATP made Electron shuttles span membrane 2 NADH Glycolysis 2 Pyruvate Glucose 2 NADH Pyruvate oxidation 2 Acetyl Co. A 2 ATP Maximum per glucose: CYTOSOL MITOCHONDRION 2 NADH or 2 FADH 2 6 NADH 2 FADH 2 Citric acid cycle Oxidative phosphorylation: electron transport and chemiosmosis 2 ATP about 26 or 28 ATP About 30 or 32 ATP
• Fermentation consists of glycolysis plus reactions that regenerate NAD +, which can be reused by glycolysis • Two common types are alcohol fermentation and lactic acid fermentation • in alcohol fermentation - pyruvate is converted to ethanol in two steps, with the first releasing CO 2 – acetylaldehyde ethanol is catalyzed by Alcohol Dehydrogenase 1 – Alcohol fermentation by yeast is used in brewing, winemaking, and baking – humans can take the ethanol and convert it back into a sugar using Alcohol Dehydrogenase Acetylaldehyde 2 ADP 2 P i Glucose 2 ADP 2 P i 2 ATP Glycolysis Glucose 2 ATP Glycolysis 2 Pyruvate 2 NAD 2 Ethanol (a) Alcohol fermentation 2 NADH 2 NAD 2 CO 2 2 Acetaldehyde 2 NADH 2 Lactate (b) Lactic acid fermentation 2 Pyruvate
• in lactic acid fermentation - pyruvate is reduced to NADH, forming lactate as an end product, with no release of CO 2 • Lactic acid fermentation by some fungi and bacteria is used to make cheese and yogurt • Human muscle cells use lactic acid fermentation to generate ATP when O 2 is scarce 2 ADP 2 P i Glucose 2 ADP 2 P i 2 ATP Glycolysis Glucose 2 ATP Glycolysis 2 Pyruvate 2 NAD 2 Ethanol (a) Alcohol fermentation 2 NADH 2 NAD 2 CO 2 2 Acetaldehyde 2 NADH 2 Lactate (b) Lactic acid fermentation 2 Pyruvate
Comparing Fermentation with Anaerobic and Aerobic Respiration • all three pathways use glycolysis (net ATP = 2) to oxidize glucose and harvest chemical energy of food • in all three - NAD+ is the oxidizing agent that accepts electrons during glycolysis (i. e. that gets reduced) • BUT these processes have different final electron acceptors: – aerobic respiration – O 2 – anaerobic respiration – other atoms – fermentation - pyruvate or acetaldehyde • Cellular respiration produces 32 ATP per glucose molecule; fermentation produces 2 ATP per glucose molecule
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