Cellular Respiration Harvesting Chemical Energy

1. Cellular RespirationHarvesting Chemical Energy ATP

2. What’s thepoint? The pointis to makeATP! ATP

3. The energy needs of life • Organisms are endergonic systems • What do we need energy for? • synthesis • building biomolecules • reproduction • movement • active transport • temperature regulation

4. Where do we get the energy from? • Work of life is done by energy coupling • use exergonic (catabolic) reactions to fuel endergonic (anabolic) reactions digestion energy + + synthesis energy + +

5. O– O– O– O– O– O– O– O– P P P P P P P P –O –O –O O– O– O– –O –O –O O– O– O– –O –O O– O– O O O O O O O O How does ATP store energy? I thinkhe’s a bitunstable…don’t you? • Each negative PO4 more difficult to add • a lot of stored energy in each bond • most energy stored in 3rd Pi • 3rd Pi is hardest group to keep bonded to molecule • Bonding of negative Pi groups is unstable • spring-loaded • Pi groups “pop” off easily & release energy AMP ADP ATP Instability of its P bonds makes ATP an excellent energy donor

6. O– O– O– O– P P P P –O O– –O O– –O –O O– O– O O O O How does ATP transfer energy? 7.3energy • ATP  ADP • releases energy • ∆G = -7.3 kcal/mole • Fuel other reactions • Phosphorylation • released Pi can transfer to other molecules • destabilizing the other molecules • enzyme that phosphorylates = “kinase” + ADP ATP

7. Harvesting stored energy • Energy is stored in organic molecules • carbohydrates, fats, proteins • Heterotrophs eat these organic molecules  food • digest organic molecules to get… • raw materials for synthesis • fuels for energy • controlled release of energy • “burning” fuels in a series of step-by-step enzyme-controlled reactions

8. glucose + oxygen  energy + water + carbon dioxide respiration ATP + 6H2O + 6CO2 + heat  C6H12O6 + 6O2 COMBUSTION = making a lot of heat energy by burning fuels in one step ATP glucose O2 O2 fuel(carbohydrates) Harvesting stored energy • Glucose is the model • catabolism of glucose to produce ATP RESPIRATION = making ATP (& some heat)by burning fuels in many small steps ATP enzymes CO2 + H2O + heat CO2 + H2O + ATP (+ heat)

9. + + oxidation reduction e- How do we harvest energy from fuels? • Digest large molecules into smaller ones • break bonds & move electrons from one molecule to another • as electrons move they “carry energy” with them • that energy is stored in another bond, released as heat or harvested to make ATP loses e- gains e- oxidized reduced + – e- e- redox

10. e p loses e- gains e- oxidized reduced + – + + H oxidation reduction H  C6H12O6 + 6O2 6CO2 + 6H2O + ATP H How do we move electrons in biology? • Moving electrons in living systems • electrons cannot move alone in cells • electrons move as part of H atom • move H = move electrons oxidation reduction e-

11. oxidation  C6H12O6 + 6O2 6CO2 + 6H2O + ATP reduction Coupling oxidation & reduction • REDOX reactions in respiration • release energy as breakdown organic molecules • break C-C bonds • strip off electrons from C-H bonds by removing H atoms • C6H12O6CO2 =thefuel has been oxidized • electrons attracted to more electronegative atoms • in biology, the most electronegative atom? • O2H2O =oxygen has been reduced • couple REDOX reactions & use the released energy to synthesize ATP O2

12. like $$in the bank O– O– O– O– P P P P –O –O –O –O O– O– O– O– O O O O NAD+ nicotinamide Vitamin B3 niacin O O H H C C NH2 C C NH2 How efficient! Build once,use many ways N+ N+ reduction + H oxidation phosphates adenine ribose sugar Moving electrons in respiration • Electron carriers move electrons by shuttling H atoms around • NAD+NADH (reduced) • FAD+2FADH2 (reduced) reducing power! NADH H carries electrons as a reduced molecule

13.  C6H12O6 + 6O2 ATP + 6H2O + 6CO2 Overview of cellular respiration • 4 metabolic stages • Anaerobic respiration 1. Glycolysis • respiration without O2 • in cytosol • Aerobic respiration • respiration using O2 • in mitochondria 2. Pyruvate oxidation 3. Krebs cycle 4. Electron transport chain (+ heat)

14. Cellular RespirationStage 1:Glycolysis

15. glucose      pyruvate 6C 3C 2x Glycolysis • Breaking down glucose • “glyco – lysis” (splitting sugar) • ancient pathway which harvests energy • where energy transfer first evolved • transfer energy from organic molecules to ATP • still is starting point for ALL cellular respiration • but it’s inefficient • generate only2 ATP for every 1 glucose • occurs in cytosol In thecytosol?Why doesthat makeevolutionarysense? That’s not enoughATP for me!

16. Evolutionary perspective Enzymesof glycolysis are“well-conserved” • Prokaryotes • first cells had no organelles • Anaerobic atmosphere • life on Earth first evolved withoutfree oxygen (O2) in atmosphere • energy had to be captured from organic molecules in absence of O2 • Prokaryotes that evolved glycolysis are ancestors of all modern life • ALL cells still utilize glycolysis You meanwe’re related?Do I have to invitethem over for the holidays?

17. enzyme enzyme enzyme enzyme enzyme enzyme enzyme enzyme ATP ATP 4 2 4 2 2 ADP NAD+ ADP 2Pi 2 2H 2Pi Overview glucose C-C-C-C-C-C 10 reactions • convert glucose (6C)to 2 pyruvate (3C) • produces:4 ATP & 2 NADH • consumes:2 ATP • net yield:2 ATP & 2 NADH fructose-1,6bP P-C-C-C-C-C-C-P DHAP P-C-C-C G3P C-C-C-P pyruvate C-C-C DHAP = dihydroxyacetone phosphate G3P = glyceraldehyde-3-phosphate

18. 2nd half of glycolysis(5 reactions) 1st Phosphoralation (adding of phosphate onto Glucose then later giving the phosphates to ADP. DHAP P-C-C-C G3P C-C-C-P • NADH production • G3P donates H • oxidizes the sugar (gets rid of H we-) • reduces NAD+ • Adds H with e- • NAD+NADH • ATP production • G3P pyruvate • PEP sugar donates P • “substrate level phosphorylation” • ADP ATP Pi Pi NAD+ NAD+ 6 NADH NADH 7 ADP ADP Energy Harvest O- phosphoglycerate kinase C ATP ATP CHOH 3-Phosphoglycerate (3PG) 3-Phosphoglycerate (3PG) CH2 P O 8 O- phosphoglycero-mutase O C H C O P 2-Phosphoglycerate (2PG) 2-Phosphoglycerate (2PG) CH2OH O- 9 H2O H2O enolase C O O C P Phosphoenolpyruvate (PEP) Phosphoenolpyruvate (PEP) CH2 O- 10 ADP ADP C O pyruvate kinase ATP ATP C O CH3 Pyruvate Pyruvate

19. 2 ATP 2 ADP 2 NAD+ 4 ADP ATP 4 2 Energy accounting of glycolysis • Net gain = 2 ATP + 2 NADH • some energy investment (-2 ATP) • small energy return (4 ATP + 2 NADH) • 1 6C sugar 2 3C sugars glucose      pyruvate 6C 3C 2x All that work! And that’s all I get? Butglucose hasso much moreto give!

20. O2 O2 O2 O2 O2 3C 2x Is that all there is? • Not a lot of energy… • for 1 billon years+ this is how life on Earth survived • no O2 = slow growth, slow reproduction • only harvest 3.5% of energy stored in glucose • more carbons to strip off = more energy to harvest glucose     pyruvate 6C Hard wayto makea living!

21. recycleNADH How is NADH recycled to NAD+? without oxygen anaerobic respiration “fermentation” with oxygen aerobic respiration Another molecule must accept H from NADH pyruvate NAD+ H2O CO2 NADH NADH O2 acetaldehyde NADH acetyl-CoA NAD+ NAD+ lactate lactic acidfermentation which path you use depends on who you are… Krebs cycle ethanol alcoholfermentation

22. [ ] 2x pyruvate  acetyl CoA + CO2 NAD Oxidation of pyruvate • Pyruvate enters mitochondrial matrix • 3 step oxidation process • releases 2 CO2(count the carbons!) • reduces 2NAD  2 NADH (moves e-) • produces 2acetyl CoA • Acetyl CoA enters Krebs cycle 1C 3C 2C Wheredoes theCO2 go? Exhale!

23. pyruvate  ethanol + CO2 3C 2C 1C pyruvate  lactic acid NADH NADH NAD+ NAD+ 3C 3C Fermentation(anaerobic) • Bacteria, yeast back to glycolysis • beer, wine, bread • Animals, some fungi back to glycolysis • cheese, anaerobic exercise (no O2)

24. pyruvate  ethanol + CO2 3C 2C 1C NADH NAD+ recycleNADH Alcohol Fermentation bacteria yeast back to glycolysis • Dead end process • at ~12% ethanol, kills yeast • can’t reverse the reaction Count thecarbons!

25. O2 pyruvate  lactic acid NADH NAD+ 3C 3C recycleNADH animalssome fungi Lactic Acid Fermentation  back to glycolysis • Reversible process • once O2 is available, lactate is converted back to pyruvate by the liver Count thecarbons!

26. O2 O2 Pyruvate is a branching point Pyruvate fermentation anaerobicrespiration mitochondria Krebs cycle aerobic respiration

27. Cellular respiration

28. outer membrane intermembrane space inner membrane cristae matrix mitochondrialDNA Mitochondria — Structure • Double membrane energy harvesting organelle • smooth outer membrane • highly folded inner membrane • cristae • intermembrane space • fluid-filled space between membranes • matrix • inner fluid-filled space • DNA, ribosomes • enzymes • free in matrix & membrane-bound What cells would have a lot of mitochondria?

29. [ ] 2x pyruvate  acetyl CoA + CO2 NAD Oxidation of pyruvate • Pyruvate enters mitochondrial matrix • 3 step oxidation process • releases 2 CO2(count the carbons!) • reduces 2NAD  2 NADH (moves e-) • produces 2acetyl CoA • Acetyl CoA enters Krebs cycle 1C 3C 2C Wheredoes theCO2 go? Exhale!

30. NAD+ 2 x [ ] Pyruvate oxidized to Acetyl CoA reduction Acetyl CoA Coenzyme A CO2 Pyruvate C-C C-C-C oxidation Yield = 2C sugar + NADH + CO2

31. Krebs cycle 1937 | 1953 • aka Citric Acid Cycle • in mitochondrial matrix • 8 step pathway • each catalyzed by specific enzyme • step-wise catabolism of 6C citrate molecule • Evolved later than glycolysis • does that make evolutionary sense? • bacteria 3.5 billion years ago (glycolysis) • free O22.7 billion years ago (photosynthesis) • eukaryotes 1.5 billion years ago (aerobic respiration = organelles  mitochondria) Hans Krebs 1900-1981

32. 2C 6C 5C 4C 3C 4C 6C 4C 4C 4C NADH ATP CO2 CO2 CO2 NADH NADH FADH2 NADH Count the electron carriers! pyruvate acetyl CoA citrate reductionof electroncarriers This happens twice for each glucose molecule x2

33. Whassup? So we fully oxidized glucose C6H12O6  CO2 & ended up with 4 ATP! What’s the point?

34. H+ H+ H+ H+ H+ H+ H+ H+ H+ Electron Carriers = Hydrogen Carriers • Krebs cycle produces large quantities of electron carriers • NADH • FADH2 • go to Electron Transport Chain! ADP+ Pi ATP What’s so important about electron carriers?

35. Cellular RespirationStage 4: Electron Transport Chain

36. Cellular respiration

37. ATP accounting so far… • Glycolysis 2ATP • Kreb’s cycle 2ATP • Life takes a lot of energy to run, need to extract more energy than 4 ATP! There’s got to be a better way! I need a lotmore ATP! A working muscle recycles over 10 million ATPs per second

38. O2 There is a better way! • Electron Transport Chain • series of proteins built into inner mitochondrial membrane • along cristae • transport proteins& enzymes • transport of electrons down ETC linked to pumping of H+ to create H+ gradient • yields ~36 ATP from 1 glucose! • only in presence of O2 (aerobic respiration) Thatsounds morelike it!

39. Mitochondria • Double membrane • outer membrane • inner membrane • highly folded cristae • enzymes & transport proteins • intermembrane space • fluid-filled space between membranes Oooooh!Form fits function!

40. Innermitochondrialmembrane Electron Transport Chain Intermembrane space C Q cytochromebc complex cytochrome coxidase complex NADH dehydrogenase Mitochondrial matrix

41. e p 1 2 Electron Transport Chain Building proton gradient! NADH  NAD+ + H intermembranespace H+ H+ H+ innermitochondrialmembrane H  e- + H+ C e– Q e– H e– FADH2 FAD H NADH 2H+ + O2 H2O NAD+ cytochromebc complex cytochrome coxidase complex NADH dehydrogenase mitochondrialmatrix What powers the proton (H+) pumps?…

42. H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ C e– Q e– 1 2 e– FADH2 FAD NADH 2H+ + O2 H2O NAD+ cytochromebc complex NADH dehydrogenase cytochrome coxidase complex Stripping H from Electron Carriers • Electron carriers pass electrons & H+ to ETC • H cleaved off NADH & FADH2 • electrons stripped from H atoms  H+ (protons) • electrons passed from one electron carrier to next in mitochondrial membrane (ETC) • flowing electrons = energy to do work • transport proteins in membrane pump H+ (protons) across inner membrane to intermembrane space H+ H+ H+ TA-DA!! Moving electronsdo the work! ADP+ Pi ATP

43. H2O O2 But what “pulls” the electrons down the ETC? electronsflow downhill to O2 oxidative phosphorylation

44. Electrons flow downhill • Electrons move in steps from carrier to carrier downhill to oxygen • each carrier more electronegative • controlled oxidation • controlled release of energy make ATPinstead offire!

45. H+ H+ H+ H+ H+ H+ H+ H+ ADP + Pi H+ We did it! “proton-motive” force • Set up a H+ gradient • Allow the protonsto flow through ATP synthase • Synthesizes ATP ADP + PiATP ATP Are wethere yet?

46. ATP Synthase Video

47. Chemiosmosis • The diffusion of ions across a membrane • build up of proton gradient just so H+ could flow through ATP synthase enzyme to build ATP Chemiosmosis links the Electron Transport Chain to ATP synthesis So that’sthe point!

48. Intermembrane space Pyruvate from cytoplasm Inner mitochondrial membrane H+ H+ Electron transport system C Q NADH e- H+ 2. Electrons provide energy to pump protons across the membrane. 1. Electrons are harvested and carried to the transport system. e- Acetyl-CoA NADH e- H2O e- Krebs cycle 3. Oxygen joins with protons to form water. 1 FADH2 O2 2 O2 + 2H+ H+ CO2 ATP H+ ATP ATP 4. Protons diffuse back indown their concentrationgradient, driving the synthesis of ATP. ATP synthase Mitochondrial matrix

50. H+ H+ H+ H+ H+ H+ H+ H+ + P H+ And how do we do that? • ATP synthase • set up a H+ gradient • allow H+ to flow through ATP synthase • powers bonding of Pi to ADP ADP + PiATP ADP ATP But… Have we done that yet?