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Home , Cellular respiration, Chemiosmosis, Intermembrane space

Lecture 15 Lecture Outline III. Oxidative + 1. What do we do with NADH + H and FADH2 reducing equivalents? 2. Transport – the oxidation phase of Oxidative Phosphorylation 3. ATP synthesis – the Phosphorylation Phase of Oxidative Phosphorylation 5. Why believe the Chemiosmotic hypothesis? 6. Indirect into the 7. Comparison of yield from Fatty Oxidation and Monosaccharide Oxidation 8. without -

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 for synthesis of ATP Energy 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 p 2 o

Free energy, G r t ATP

Given to c h

a

i Electron Transport n Generate >90% of Typical ’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 4 Figure 9.5 B (b) Cellular respiration Electron transport Oxidative part 3 discrete 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 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 energy of oxidation pumps H to intermembrane

++ space 2 Fe 2e- and 2H+ (where from?) get passed to another complex (complex III) + - energyorphaning of oxidation of 2H pumpsrepeated H as 2e passed on - + to (2e ) to intermembrane 2 Fe+++ (2e-) (2e-) CoQ CoQ - + space 2e- + 2e and 2H (where from?) get passed to a third complex (spacecomplex IV) 2H energyorphaning of oxidation of 2H+ repeatedpumps H as 2e- passed on + 2H (oxidized) H H finally to oxygen + to to intermembrane pH 8 (reduced) + + NADH + H NAD 7 space 8 need to pick up + (where from?)to form water 2H + 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+ ?

+ 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 to ADP pH 8 i pH 8 to make ATP. 13 14 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 phosphorylation. electron transport 2 ATP forand chemiosmosiseach FADH2

ATP ATP ATP H+ • and theH+ electron transport chain H+ H+ •Agents such as poison electron transport Cyt c 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 + 2 H+ + 1/ O diminish ATP synthesis membrane FAD 2 2 NADH+ + NAD ADP + P i ATP + •Agents that allow H passage (dissipate gradient) (Carrying electrons from, food) H+ Mitochondrial Chemiosmosis – matrix Electron transport chain dinitrophenol Electron transport and pumping of (H+), ATP synthesis powered by the flow which create an H+ gradient across the membrane Of H+ back across the membrane

(DNP) 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 oxidation? Electrochemical Gradient ~36 ATP per 6 carbon 7.3 kcal/mole x 36 = 263 kcal/mole Can also power ∆G = -686 kcal/mole 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 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 Fatty acid

18 carbon fatty acid (9 two carbon units) In : Each 2 carbon unit 1 X2 NADH + H+ 3 6 ATP X Highly reduced fully oxidized 1 X2 FADH2 2 X4 ATP X (captured) CH Each acetyl CoA gives in TCA 3-CH2-CH2-(CH2)x-CH2-C-O + O2 H2O + CO2 + energy 3 NADH + H+ 9 ATP 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 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 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 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 Fermentation Cellular respiration Method CO Wine/beer To Recycle - + 2 NADH 2e 2 NAD X CH + 3 NADH + H + O MITOCHONDRION 2H Acetyl CoA Recycles CO or lactate NADH H C OH CH Muscles cycle Keeps the 3 Run in reverse two2 Lactate When oxygen 29 Motor running 30 (b) 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 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 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. Oxidative Phosphorylation is comprised of two distinct processes Electron Transport (oxidative) makes H+ gradient Next time: ATP synthesis (phosphorylation) uses H+ gradient coupled by H+ gradient

Light energy 2. Processes are separable: cyanide (poisons electron transport) ECOSYSTEM uncouplers (DNP) (dissipate H+ gradient)

Photosynthesis 3. Electrochemical gradient powers in Organic CO + H O + O + 2 2 Cellular 2 mitochondrial H ATPase respiration in mitochondria indirect active transport

4. Oxidation of fatty acids yields more ATP than oxidation of glucose ATP powers most cellular work 5. Can ferment in absence of oxygen Heat still produce ATP by substrate level phosphorylation Figure 9.2 energy 33 but no reducing equivalents captured 34 Why not?

• The catabolism of various molecules from food

Proteins

Amino Sugars Fatty acids acids

Glycolysis Glucose

Glyceraldehyde-3- P

NH3 Pyruvate

Acetyl CoA

Citric acid cycle

Oxidative Figure 9.19 phosphorylation 35