1. Electron Transport and ATP Synthesis C483 Spring 2013

2. 1. To reduce one molecule of O2, ________ electron(s) must be passed through the electron transport chain and ________ molecule(s) of NADH is(are) oxidized. A) 4; 2 B) 2; 1 C) 1; 1 D) 1; 2 E) 4; 4 2. The chemiosmotic theory is a concept that ________. A) the transport of Na+ and K+ across cell membranes is by active transport B) explains how transport by facilitated diffusion reaches a saturation limit C) explains the blood-brain barrier D) a proton gradient drives the formation of ATP 3. Which is the proper ranking of greatest to least reduction potential? A) NAD+, Q, O2 B) O2, Q, NAD+ C) Q, NAD+, O2 D) Q, O2, NAD+

3. 4. According to the binding change mechanism, the (alpha3 beta3 ) oligomer of ATP synthase has 3 catalytic sites which can each have ________ different conformations. A) 2 B) 3 C) 6 D) 9 5. The P/O ratio for passing electrons through complexes I, III and IV is ________. A) 1 B) 1.5 C) 2 D) 2.5 E) 3

4. Goal: ATP Synthesis

5. Overview Redox reactions Electron transport chain Proton gradient ATP synthesis Shuttles Analogy: How does burning coal put flour in the grocery store?

6. Redox reactions: electricity 2 e- transfer Calculate G by reduction potential NADH: E o = -.32 FMN: E o =-.30 G o = -nF E o = -2(96485)(0.02) = -3.9 kJ/mol

7. Coenzyme Q: Mobile Carrier FADH 2 is a one e- donator Many reactions, including metals Ubiquinone is a key intermediate Can diffuse through nonpolar regions easily

8. Numerous Redox Substrates O 2 : high reduction potential Substrates – Organic cofactors – Metals (iron/sulfur clusters) – cytochromes

9. Oxygen: the final electron acceptor Water is producedhas very low reactivity, very stable Superoxide, peroxide as toxic intermediates Overall reaction NADH + H + + ½ O 2 NAD + + H 2 O

10. Flow Through Complexes

11. Downhill Flow of Electrons

12. Compartmentalization

13. Protonmotive Force Flow of electrons is useless if not coupled to a useful process – Battery connected to wire Proton gradient across mitochondrial membrane

14. Overview of Complexes I-IV Dont need to know which cofactors in which complexes, mechanism of proton pumping Complex I and II are different entry points into Q pool, which goes to Complex III

15. Protonmotive Force NADH + H + + ½ O 2 NAD + + H 2 O + 10 H + pumped succinate + ½ O 2 fumarate + H 2 O + 6 H + pumped

16. Proton Gradient Gradient driven by concentration difference + charge difference Free energy of ATP hydrolysis = -48 kJ/mole How many protons needed to fuel ATP formation? Minimum of 3

17. Using the Gradient Coupled to ATP synthesis Uncouplers used to show link of oxygen uptake and ATP synthesis

18. Complex V: ATP Synthase Molecular motor Rotor: c,, – Proton channel

19. Proton Channel Protons enters channel between rotor and stator (unit a--purple) Rotor rotates to release strain by allowing proton to enter matrix Stalk ( ) moves inside the knobhexameric ATP synthase 9 or 10 protons = full rotation

20. Binding-Change Mechanism Stalk causes ATP synthase to have three different conformations: open, loose, tight In tight conformation, energy has been used to cause an energy conformation that favors ATP formation 9 protons = 3 ATP (or 1 ATP/3 protons)

21. Remember Analogy Fuel electricity water pumped uphill flows down to grind flour But we dont have bread until flour is transported to where it needs to go! Compartmentalization: ATP is in matrix, but must get to the rest of the cell

22. Active Transport of ATP ATP must go out, ADP and P i must go in Together, use about 1 proton of protonmotive force

23. Energy Accounting ATP costs 4 protons – 3 protons in ATP synthase, 1 proton in transport NADH pumps 10 protons – 4 protons in Complex I, 4 protons in Complex III, and 2 protons in Complex IV – 2.5 ATP/NADH called P/O ratio--# of phosphorylation per oxygen atom QH 2 pumps 6 protons – 4 protons in Complex III and 2 protons in Complex IV – 1.5 ATP/QH 2

24. Net ATP Harvest from Glucose Glycolysis = 2 ATP – Plus 3 or 5 ATP from NADH – What leads to difference in this case? Pyruvate DH = 5 ATP Citric Acid Cycle = 20 ATP Total: 30-32 ATP/glucose

25. NADH Shuttles Glycerol phosphate shuttle (1.5 ATP/NADH) Produces QH 2 Operational in some tissues/circumstances

26. NADH Shuttles Malate-Aspartate shuttle (2.5 ATP/NADH) – Actually slightly less because one proton is consumed in shuttle Separate NAD+/NADH pools indirectly interconverted Operational in some tissues/circumstances

27. Answers 1.A 2.D 3.B 4.B 5.D