MMG 301, Lecture 20 RespirationQuestions for today:1. What is Respiration(and how does it differ from fermentation)?2. What substrates are oxidizedand which reduced(aerobic vs anaerobic respiration)? 3. How are electrons transferredfrom the electron donor to the electron acceptor?5. How is ATPgenerated?4. What are examplesof respiration?6. How do we calculate the energyavailable from respiration?Overview of Respiration-- Twosubstrates are required (A & B)-- One substrate (A; organic or inorganic compound) is oxidized-- The second substrate (B; O2, nitrate, sulfate, or other compound) is reduced. This is referred to as the final electron acceptor -- Electron transfer reactions drive ATP synthesis by “Electron Transport Phosphorylation” (ETP), little if any SLPA →→→→→ Pe-B →→ QChemoorganotrophic Respiration: an organic compound is oxidized (typically to CO2) and the electrons transferred to oxygen or another acceptorChemolithotrophic Respiration: an inorganic compound is oxidized and the electrons transferred to oxygen or another acceptorNote the clear differences from the situation in fermentation!Examples of Substrate OxidationA. PyruvateoxidationMany organic compounds are converted to pyruvate(e.g., glycolysis). Each pyruvate is oxidized to 3 CO2by the Citric Acid CycleElectrons reduce 4 NAD + 1 FAD to form 4 NADH and 1 FADH2that are used for ETP 1 GTP made by SLP (used to form 1 ATP)B. Acetyl-CoA oxidationVarious organic compounds (including fatty acids) are decomposed to → acetyl-CoA Oxidized via Citric Acid Cyclewith electrons going to 3 NAD and 1 FADC. Non-Citric Acid Cycleoxidation of organiccompoundsExample: Respiration of single carbon compounds:CH4→→→→ CO2multiple electrons (used for ETP)D. Oxidation of InorganicsubstratesH2→ 2 H++ 2 e-(used for ETP)H2S → 2 H++ So+ 2 e-(used for ETP)NO2-→ NO3-+ e-(used for ETP)Fe2+→ Fe3++ e-(used for ETP)(each pathway for C and D will require a separate series of enzymes for the specific oxidation reactions)MethanotrophsElectron Transport ChainsElectrons derived from substrate are handed off through a series of electron carriers:NAD(P) -- shown last time; 2H+and 2e-Flavins Quinones (Q)Iron-sulfur (Fe/S) clusters: only electron transfer!You should be able to recognize these structures!Cytochromes (Cyt): only electron transfer!These cofactors (or proteins containing them) are positioned in or on the cytoplasmic membranein a manner that allows for a “bucket-brigade” transfer of electrons while transferring protons across the membrane: proton motive force (PMF)The electrons are finally used to reduce the final electron acceptor substrate: O2(aerobic respiration) or nitrate, nitrite, sulfate, fumarate, ferric ion, and other compounds (anaerobic respiration)Each pathway requires a distinct series of electron transfer proteins and a unique oxidaseExample: aerobic respiration inParacoccus denitrificans and Escherichia coliMembranes allow buildup of PMF thatdrives ATP synthesis via ATP synthasePutting it all together: examples of RespirationAerobic Respiration of Glucose-- Glucose (A) is metabolized to pyruvate via glycolyis and oxidized to CO2by the citric acid cycle.-- NADH and FADH2feed electrons into membrane bound electron carriers that result in protons being pumped.-- Electrons oxidize oxygen (B) to produce water.-- PMF drives ATP synthesis.Aerobic respiration of other carbon compounds-- Alternative pathways are used for oxidation of the carbon compound to provide reduced NAD and FAD (e.g. CH4).Aerobic Respiration in Lithotrophs occurs by an analogous system-- An inorganic substrate is oxidized (e.g., hydrogen, sulfide, nitrite, ferrous ion).-- Electrons feed through an electron transport chain to generate PMF.-- Oxygen is reduced to water.HydrogenoxidationSulfideOxidation(sulfide oxidase)Nitrite oxidation(nitrite oxidoreductase)Ferrous ion oxidationUnderstand the concept and recognize the key enzymes indicated, not the detailed pathwaysAnaerobic Respiration: something other than oxygen is reduced (e.g., nitrate, sulfate, fumarate, etc.)Denitrification:Energy-generating nitrate reduction in E. coli vs. PseudomonasNote that some portions of the electron transport chain may be identical to that used for aerobic respirationDissimilative sulfate reduction: Energy generation by anaerobic respiration of sulfateLactic acid or other carbon compound is oxidized with the electrons reducing sulfate to form sulfideOther electron acceptors used for anaerobic respiration:fumarate + 2H++ 2e-→ succinatedimethylsulfoxide + 2H++ 2e-→ dimethylsulfideferric ion + e-→ ferrous ion(many others)Only carried out by one closely related group of microorganisms: the sulfate reducersMeasurement of EnergyDirect calculation of ∆Go’Glucose + 6 O2→ 6 CO2+ 6 H2O(-917 kJ/mol) (0) (-394 kJ/mol)(-237 kJ/mol)∆Go’= [6(-394)+6(-237)]-(-917) = -2869 kJ/mol10-fold that available via fermentation!Find ~38 ATP are made (2 during glycolysis, 2 GTP from citric acid cycle, and the rest via ETP)Efficiency = 100 [38*(-32 kJ/mol)/-2869 kJ/mol) = 42%This approach also works when using different electron acceptors (I.e., types of anaerobic respiration) and when using different substrates to be oxidized (e.g., H2, H2S, NO3-, etc.). However, the values of free energies are not always readily available (see table A1.1). Is there an easier way to calculate energy for these reactions?Calculation of energy by∆Eo’(measured in V)These reactions all involve two half-reactionsdonor substrate product 1e-acceptor substrate product 2Eo’is the tendency of a compound to become reduced, in units of V∆Eo’= Eo’(electron acceptor) - Eo’(electron donor) Using hydrogen gas as the electron donor:Fumarate(.03)-(-.43)= 0.46 VNitrate(0.42)-(-.43) = 0.85 VOxygen(.82)-(-.43)= 1.25 VElectrons flow down!How do these values compare to ∆Go’?∆Go’= -nF∆Eo’F = 96.48 kJ/mol n = number of electrons So we can compare the three reactions!∆Eo’∆Go’H2+ fumarate → succinate 0.46 V ~-85 kJ/molH2+ NO3-→ NO2-+ H2O 0.85 V ~-160 kJ/molH2+ ½ O2→ H2O 1.25 V ~-240 kJ/molCan estimate the ATP available from such processes on the basis of –32 kJ/mol = ~0.16 V (for a 2 electron process)Things to think about regarding oxidation• What is needed for a cell to carry out aerobic respiration of glucose?• What components would change if the cell is carrying out nitrate respiration, a type of anaerobic respiration?• How might a cell
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