Lecture 15 Cellular Respiration Lecture Outline III. Oxidative + Phosphorylation 1. What do we do with NADH + H and FADH2 reducing equivalents? 2. Electron Transport – the oxidation phase of Oxidative Phosphorylation 3. ATP synthesis – the Phosphorylation Phase of Oxidative Phosphorylation 5. Why believe the Chemiosmotic hypothesis? 6. Indirect Active Transport into the mitochondrion 7. Comparison of yield from Fatty Acid Oxidation and Monosaccharide Oxidation 8. Life without oxygen - Fermentation October 5, 2005 Chapter 9 1 2 1 2 H + /2 O2 Mitochondrial Functions Electron transport (from food via NADH) – oxidizes NADH Controlled and FADH2 2 H+ + 2 e– release of Oxidize compounds to CO2 + H2O energy for synthesis of ATP Energy Fatty acid Oxidation E ATP + l Produce back to NAD e c Released t Released r and FAD o ATP Oxidation of Pyruvate n reduced carriers Used To reduced carriers t r a n TCA Kreb’s Cycle s ATP Make NADH & FADH Electrons p 2 o Free energy, G r t ATP Given to c h a i Electron Transport n Generate >90% of Typical Cell’s ATP chain Oxidative Phosphorylation 2 e– Oxidative 1 /2 O2 2 H+ “electron transport” Electrons ATP synthesis Oxidize reduced carriers Eventually Given to H O 2 How? to produce ATP or equiv Oxygen 3 To form Water 4 Figure 9.5 B (b) Cellular respiration Electron transport Electron Transport Chain Oxidative part 3 discrete Proton pumps Proton pumps Oxidize NADH + H+ to NAD+ Powered by 2e- and 2H+ given to a multiprotein complex (complex I) Passage of 2e- get passed on electrons 2H+ orphaned as electrons transferred energy of oxidation pumps H+ to intermembrane space Need O2 as final acceptor of reducing equivalents 5 6 2H2 + O2 = H2O + BOOM! Electron Transport Chain Complex I pH 6 close up Oxidize NADH + H+ to NAD+ 2e- and 2H+ given to multiprotein complex (complex I) 2e- get passed on 2H+ 2H+ orphaned as electrons transferred energy of oxidation pumps H+ to intermembrane space ++ 2 Fe 2e- and 2H+ (where from?) get passed to another complex (complex III) orphaning of 2H+ repeated as 2e- passed on (2e-) energy of oxidation pumps H+ to intermembrane space 2 Fe+++ (2e-) (2e-) CoQ CoQ - + 2e- + 2e and 2H (where from?) get passed to a third complex (complex IV) 2H orphaning of 2H+ repeated as 2e- passed on + 2H (oxidized) H H finally to oxygen energy of oxidation pumps H+ to intermembrane space pH 8 (reduced) + + NADH + H NAD 7 8 need to pick up 2H+ (where from?)to form water + NADH H 50 2 What happened to FADH pH 6 2 complex II? I Multiprotein 40 FMN FAD complexes II Fe•S Fe•S O III + Cyt b Three 2H pumping (kcl/mol) Fe•S 2 30 + Cyt c Steps from NADH + H 1 IV oxidation Cyt c oxidation ) relative to O Cyt a 2 G Cyt a3 20 + Only TWO 2H pumping Free energy ( Steps from FADH pH 8 OH- Steps from FADH2 oxidation 10 Net Result of Electron Transport: 0 2 H + + 1⁄ O2 an electrochemical H+ gradient 2 9 10 Figure 9.13 H2O What good is the H+ Electrochemical Gradient ? + Electron transport – oxidizes NADH and FADH2 H back to NAD+ and FAD OH- H O Electron transport – electrons passed in series 2 from one carrier to another pH 6 eventually give e- and H+ to oxygen to form water + BUT PUMP H in the process across the inner mitochondrial membrane into intermembrane space pH 8 FORMS BIG H+ gradient 11 12 INTERMEMBRANE SPACE pH 6 + Mitochondrial H A rotor within the pH 6 Mitochondrial + H H+ membrane spins clockwise when + H+ flows past H+ H ATPase H+ it down the H+ gradient. H+ H+ ATP synthase A stator anchored in the membrane holds the knob stationary. A rod (for “stalk”) extending into the knob also + spins, activating H movement catalytic sites in the knob. drives H+ conformational Three catalytic change ADP sites in the stationary knob pH 8 + join inorganic P ATP Phosphate to ADP pH 8 i pH 8 to make ATP. 13 14 MITOCHONDRIAL MATRIX Figure 9.14 Mitochondrial H+ ATPase Reducing Electron equivalents Transport ATP power the pump ADP + Pi Chain is a H+ Proton movt Pump H+ H+ Open ATP Open ADP + Pi ATP Gradient powers expelled15 16 ATP Synthesis Uncoupling Oxidation and Phosphorylation Roughly + Inner •Must have intact membranes to make ATP 3 ATP for each NADH + H Mitochondrial Oxidative membrane Glycolysis phosphorylation. electron transport 2 ATP forand chemiosmosiseach FADH2 ATP ATP ATP H+ • Chemiosmosis and theH+ electron transport chain H+ H+ •Agents such as cyanide poison electron transport Cyt c Protein complex chain Intermembrane of electron so no H+ gradient produced space carners Q IV -but if supply H+ gradient can still make ATP I III ATP Inner II synthase mitochondrial FADH 2 H2O + + 1 membrane FAD 2 H + /2 O2 NADH+ + NAD ADP + P i ATP + •Agents that allow H passage (dissipate gradient) (Carrying electrons diminish ATP synthesis – dinitrophenol (DNP) from, food) H+ diminish ATP synthesis – dinitrophenol (DNP) Mitochondrial Chemiosmosis matrix Electron transport chain Electron transport and pumping of protons (H+), ATP synthesis powered by the flow which create an H+ gradient across the membrane Of H+ back across the membrane Figure 9.15 Oxidative phosphorylation 17 18 H+ pH 8 ∆ pH 6 ∆pH drives Can Use H+ gradient to power -OH pyruvate + other useful work H import H+ -OH How you get pyruvate- (an acid) across the H+ H+ mitochondrion membrane? (not magic!) -OH ∆Ψ H+ ∆Ψ (charge) drives -- - How about ADP-- and P - into mitochondrion? ATP/ADP ADP Pi H+ antiport H+ -OH --- H+ How about ATP out of mitochondrion? ∆pH drives - Pi import 19 OH 20 So how many ATP from Glucose oxidation? Electrochemical Gradient ~36 ATP per 6 carbon sugar 7.3 kcal/mole x 36 = 263 kcal/mole Can also power ∆G = -686 kcal/mole CYTOSOL Electron shuttles MITOCHONDRION span membrane 2 NADH ~39% efficient or 2 FADH Indirect ACTIVE TRANSPORT 2 2 NADH 2 NADH 6 NADH 2 FADH2 Glycolysis Oxidative 2 Citric of IONS into MATRIX 2 phosphorylation: Acetyl acid Glucose Pyruvate electron transport CoA cycle and chemiosmosis instead of making ATP + 2 ATP + 2 ATP + about 32 or 34 ATP by substrate-level by substrate-level by oxidative phosphorylation, depending phosphorylation phosphorylation on which shuttle transports electrons from NADH in cytosol About Maximum per glucose: 36 or 38 ATP 21 22 Figure 9.16 Compare to Fatty Acid Oxidation 18 carbon fatty acid (9 two carbon units) In Metabolism: Each 2 carbon unit 1 X2 NADH + H+ 3 X6 ATP 1 2 FADH Highly reduced fully oxidized 1 X2 FADH2 2 X4 ATP CH Each acetyl CoA gives in TCA 3-CH2-CH2-(CH2)x-CH2-C-O + O2 H2O + CO2 + energy 3 NADH + H+ 9 ATP Fatty acid O (captured) 2 ATP 1 FADH2 1 GTP 1 ATP Partially reduced fully oxidized Each 2 carbons 17 22 ATP Each 2 carbons 17 22X ATP + O2 H2O + CO2+ energy 153 Oxidation of 18 carbon fatty acid yields 198X153 ATPATP carbohydrate (captured) 8.5 X11 ATP per FA carbon Why? 6 ATP per glucose carbon 23 24 Organic Oxidation Series Life Without Oxygen CH4 Inner Mitochondrial Oxidative membrane Glycolysis phosphorylation. R-CH -CH electron transport 2 3 Fatty Acids and chemiosmosis ATP ATP ATP Start Here H+ R-CH=CH + 2 • Chemiosmosis and theH electron transport chain + X H H+ Cyt c Protein complex R-CH2-CH2 -OH Intermembrane of electron space carners Monosaccharides Q IV R-CH -C=O I XIII 2 Start Here ATP Inner FADHII synthase mitochondrial FADH2 H 2 H2O + + 1 membrane X FAD 2 H + /2 O2 NADHNADH+ X NAD+ + X ADP + PX i ATP + H X R-CH2-C=O (Carrying electrons from, food) H+ Mitochondrial Chemiosmosis OH matrix Electron transport chain Electron transport and pumping of protons (H+), ATP synthesis powered by the flow which create an H+ gradient across the membrane Of H+ back across the membrane Figure 9.15 Oxidative phosphorylation O=C=O 25 26 Life Without Oxygen The Regeneration Energy Carriers Energy carriers (ATP, NAD+, FAD) present Electrons Electrons carried in only minute amounts carried X via NADH and via NADH FADH2 - 2e 2e- + 2e Can’t 2H 2H+ X Captured in catabolism Cashed in Oxidative Citric Glycolsis phosphorylation: X acid Glucose Pyruvate electron cycle transport and Run out NADH + H+ chemiosmosis X X Of Cytosol + Mitochondrion NAD ATP ATP ATP Energy from catabolism Energy for cellular work (exergonic, energy yielding (endergonic, energy- Substrate-level processes) consuming processes) Substrate-level Oxidative phosphorylation X + phosphorylation phosphorylation NAD Figure 9.6 27 X 28 Life Without Oxygen - FERMENTATION P 2 ATP Glucose 2 ADP + 2 1 Running two Muscles CYTOSOL Muscles pyruvate Glucose Pyruvate Glycolysis O– No O present 2 O2 present CO Yeast Fermentation Cellular respiration Method CO Wine/beer To Recycle - + 2 NADH 2e 2 NAD X CH + 3 NADH + H + O MITOCHONDRION 2H Ethanol Acetyl CoA Recycles CO or lactate NADH H C OH Citric acid CH Muscles cycle Keeps the 3 Run in reverse two2 Lactate When oxygen 29 Motor running 30 (b) Lactic acid fermentation returns Figure 9.18 Figure 9.17 lactate P 2 ATP – Fermentation and cellular respiration 2 ADP + 2 1 O two – Differ in their final electron acceptor C O pyruvate - oxygen CO - lactate Glucose Glycolysis - or ethanol CH3 2 Pyruvate 2e- Cellular aerobic respiration + 2 NADH + 2 NAD 2 CO – Produces tons more ATP 2H 2 – Produces tons more ATP Gives the » 2 ATP in fermentation Recycles » 36 ATP in aerobic oxidation of glucose H H Fizz NADH To alcohol H C OH CO Champagne Fermentation allows organisms CH Beer 3 CH3 Hard cider to eek out a living in low or no O2 2 Ethanol 2 Acetaldehyde Figure 9.17 two 31 32 ethanol(a) Alcohol fermentation Ancient system – pre oxygen Summary: Where does oxygen come from? 1.