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1 Electron Transport System Slide 2 2 There are 2 Ways to Make ATP 1. Substrate phosphorylation 2. Electron transfer-dependent oxidative phosphorylation Slide 3 3 2 Glycolytic Reactions Make ATP by Substrate-level Phosphorylation --1,3-BPG is an energy rich molecule with a greater phosphoryl-transfer potential than that of ATP. Thus, it can be used to power the ATP synthesis from ADP. --This is called substrate-level phosphorylation because the phosphate donor is a Substrate with high phosphoryl-transfer potential. Slide 4 4 2 Glycolytic Reactions Make ATP by Substrate-level Phosphorylation PEP has high phosphoryl-transfer potential, pyruvate (ketone) is much more stable than enol form. Slide 5 5 There are 2 Ways to Make ATP 1. Substrate phosphorylation 2. Electron transfer-dependent oxidative phosphorylation Slide 6 6 How do we obtain lots of ATP? Glucose Reduced coenzymes (NADH + H +, FADH 2 ) O2O2 H2OH2O Glycolysis TCA ETC ATP Food (carbohydrates) ATP Little Lots (~4 ATP) (~28-30 ATP) After TCA cycle, energy is extracted In the form of reduced Coenzymes, FADH2 and NADH Electron transport and Oxidative phosphorylation: Involved many steps, Sequestered in special environment. Glycolysis Slide 7 7 Minimal TCA CycleGlucose Pyruvate NADH + H + CO 2 NADH + H + CO 2 GDP GTP FADH 2 NADH + H + 6C4C 4C (2C) CoASH CH 3 C-SCoA O 1 GTP 1 GTP 3 NADH 3 NADH +1 FADH 2 10 ATP/cycle And releases two CO 2 NOTE: 1 NADH 2.5 ATP; 1 FADH2 1.5 ATP; 1 GTP 1 ATP so get 1 + 7.5 + 1.5 = 10 ATP/cycle NADH + H + Slide 8 8 Where in the cell does electron transport and oxidative phosphorylation occur? Slide 9 9 Slide 10 10 Mitochondria Permeable Outer Mtch Membrane Intermembrane Space Inner Mtch Membrane Matrix TCA enzymes -oxidation ATP synthase e - transport chain M DNA Slide 11 11 Mitochondria --A mitochondrion is bounded by a double membrane, with an intermembrane space. --Outer M: permeable to most ions and small molecules --The inner membrane: highly impermeable, Highly folded cristae. most molecules require transporters (exceptions: O2, CO2). provide large surface area for the transport proteins, several FAD-dependent dehydrogenases and all enzymes and proteins of oxidative phosphorylation --The matrix is the fluid-filled interior of the mitochondrion. oxidative enzymes like pyruvate dehydrogenase (acetyl Co A formation) glutamate dehydrogenase, TCA cycle enzymes, fatty acid oxidation enzymes --Note that glycolysis occurs outside the mitochondrion in the cytosol, whereas the citric acid cycle occurs in the matrix. --The electron transport system is located on the cristae, both TCA cycle and oxidative phosphorylation occur within the mitochondrion. Slide 12 12 Electron Transport System (ETS) The electron transport system is located in the cristae of mitochondria It is a series of protein/prosthetic group carriers that pass electrons from one to the other. Electrons are donated to the ETS by NADH and FADH 2 As a pair of electrons is passed from carrier to carrier, energy is released and is used to form ATP At the end of the electron transport chain, oxygen receives the energy-spent electrons, resulting in the production of water. O 2 + 2 e- + 2 H + H 2 O (Oxygen is the final electron acceptor) Slide 13 Redox Reactions e- ++ A A B B oxidation reduction O oxidation I is L loss of electrons R reduction I is G gain of electrons Reductant (A): is oxidized, electron donor Oxidant (B): is reduced, electron acceptor Slide 14 14 How are redox potentials determined? Slide 15 15 Half cell reactions measure electromovtive force SampleReference Neg value = oxidized form has a lower affinity for electrons than does H 2 (e.g., NADH a strong reducing agent has a negative reduction potential) Pos value = oxidized form has a higher affinity for electrons than does H 2 (e.g., Oxygen a strong oxidizing agent has a positive reduction potential) Standard: 1M H+ 1atm H 2 gas E 0 of H + /H 2 is 0 volts Reductant Oxidant Ethanol gives up e to H + to form H2 H2 gives up e to Fe 3+ to form H + Slide 16 16 A strong reducing agent, NADH is poised to donate electrons, has a negative reduction potential, whereas a strong oxidizing agent O2 is ready to accept electrons and has a positive reduction potential. --Biochemists use E 0 , the value at pH 7. --Chemists use E 0, the value in 1M H +. --The prime denotes that pH 7 is the standard state. --Thus, these values are different in chem textbooks. Slide 17 17 Partial reactions By convention, reduction potentials (as in Table 18.1) refer to partial reactions are written as:Table 18.1 oxidant + e - reductant OVERALL REACTION Slide 18 18 Redox reactions Redox pairs act as e - carriers Reductant + oxidant oxidized reductant + reduced oxidant Free energy is released in the transfer of e - e- ++ A A B B oxidation reduction (OIL) (RIG) Slide 19 19 Standard free-energy changes of an oxidation- reduction reaction can be determined G 0 = -nF E 0 G 0 = standard free-energy change F= faraday constant = 23.06 kcal/mol/V (required to remember!) n = number of electrons E 0 = Change in reduction potential G 0 : standard free energy change for a redox reaction is related to the difference in E 0 between the e - acceptor and donor Slide 20 20 Determining: G 0 : standard free energy change E 0 = E 0 (acceptor) - E 0 (doner) G 0 = -nF E 0 F= faraday constant = 23.06 kcal/mol/V n = number of electrons G0G0 = -2 x 23.06 kcal/mol/V x [-0.19 (-0.32) V ] -6.0 kcal/mol Pyruvate NADH = -2 x 23.06 kcal/mol/V x 0.13V = Slide 21 21 1.14 Volt potential favors formation of proton gradient Note: G 0 = -7.3 kcal/mol for the hydrolysis of ATP Acceptor donor G 0 = -nF E 0 = -nF (E 0 acceptor E 0 donor ) = -2 x 23.06 kcal/mol/V x [0.82V- (-0.32V)] = -2 X 23.06 kcal/mol/V x 1.14V = -52.6 kcal/mol The driving force of oxi phos is the elec-trans potential of NADH or FADH2 rel. to that of O2. The released energy is used to generate a proton gradient, then for ATP synthesis Slide 22 22 Driving e - Transport Electron carriers at the beginning of the chain are more - E 0 than those at the end so e - flow spontaneously from NADH (E 0 = 0.32 v) or FADH 2 (E 0 = 0.22V) to O 2 (E 0 = +0.82 volts) Neg reduction potential = oxidized form has a lower affinity for electrons and so transfers them most easily to an acceptor Pos reduction potential = will be the strongest oxidizing substance and have a higher affinity for electrons Slide 23 23 The electron transport system consists of four protein complexes and two mobile carriers. NADH-Q Oxidoreductase Succinate-Q reductase Q-cytochrome c Oxidoreductase Cytochrome c Oxidase Coenzyme Q Cytochome c The mobile carriers transport electrons between the complexes, which also contain electron carriers. The carriers use the energy released by electrons as they move down the carriers to pump H+ from the matrix into the intermembrane space of the mitochondrion. complexes carrier Slide 24 24 NAD+/NADH Fumarate/ Succinate Cytochrome C (+3) / (+2) Slide 25 25 A very strong electrochemical gradient is established with few H+ in the matrix and many in the intermembrane space. The cristae also contain an ATP synthase complex through which hydrogen ions flow down their gradient from the intermembrane space into the matrix. The flow of three H+ through an ATP synthase complex causes a conformational change, which causes the ATP synthase to synthesize ATP from ADP + P. Slide 26 26 Mitochondria produce ATP by chemiosmosis, so called because ATP production is tied to an electrochemical gradient, namely an H+ gradient. Once formed, ATP molecules are transported out of the mitochondrial matrix. Slide 27 27 Mitchells Postulates for Oxidative Phosphorylation 1.The respiratory and photosynthetic electron transfer chains should be able to establish a proton gradient 2.The ATP synthases should use the proton-motive force to drive the phosphorylation of ADP 3.Energy-transducing membranes should be impermeable to protons. If proton conductance is established (uncouplers), a proton-motive force should not form and ATP synthesis should not occur. 4.Energy-transducing membranes should possess specific exchange carriers to permit metabolites to permeate in the presence of high membrane potential ADP ATP H+H+ H+H+ H+H+ e-e- H+H+ ADP + P i ATP Mitochondrial matrix ATP-ADP Antiporter Intermembrane