In the 1940s, some physicians prescribed the drug DNP (dinitrophenol) to help patients lose weight. This treatment was abandoned after several patients died. DNP works by uncoupling the chemiosmotic machinery by making the lipid bilayer of the inner mitochondrial membrane leaky to H*. In terms of respiration, explain how DNP can lead to weight loss.
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- 2. Mitochondria isolated from bovine cardiac muscle, when subjected to sonication, form closed mem- brane vesicles containing the electron transport catalysts and the ATP synthase enzyme located in the inner mitochondrial membrane. A diagrammatic representation of these electron transport particles retaining the capacity for oxidative phosphorylation is given below. NADH NAD+ H₂O tion? 1/202 Q₁H₂ NADH- NAD+ Complex I Complex III ADP Complex IV Complex II ATP ADP ATP In the diagram protein complexes colored gray indicate that they are inhibited while complexes colored red, green, etc. are catalytically active. (a) In Panel A indicate which side of the membrane vesicle faced the matrix and which side faced the intermembrane space prior to sonication. During coupled substrate oxidation which compart- ment will be of higher pH and which of lower pH. (b) For the system in Panel A, sonication of mitochondria was carried out in the presence of an excess of ferricytochrome c (i.e., the heme…2. Mitochondria isolated from bovine cardiac muscle, when subjected to sonication, form closed mem- brane vesicles containing the electron transport catalysts and the ATP synthase enzyme located in the inner mitochondrial membrane. A diagrammatic representation of these electron transport particles retaining the capacity for oxidative phosphorylation is given below. NADH NAD+ ( ATP tion? H₂O ADP 1/202 NADH Q₁H₂ NAD+ Q₁ Complex I Complex III ADP Complex IV ATP In the diagram protein complexes colored gray indicate that they are inhibited while complexes colored red, green, etc. are catalytically active. Complex II (a)( In Panel A indicate which side of the membrane vesicle faced the matrix and which side faced the intermembrane space prior to sonication. During coupled substrate oxidation which compart- ment will be of higher pH and which of lower pH. (b) For the system in Panel A, sonication of mitochondria was carried out in the presence of an excess of ferricytochrome c (i.e., the…In the 1930s, some physicians prescribed low doses of a compound called dinitrophenol (DNP) to help patients lose weight. This unsafe method was abandoned after some patients died. DNP uncouples the chemiosmotic machinery by making the lipid bilayer of the inner mitochondrial membrane leaky to H+ . Explain how this could cause weight loss and death.
- 3. The free energy associated with the proton gradient that develops across the inner mitochondrial membrane as a result of the electron transport chain is 23.3 kJ per mole of protons. If FADH; is the only electron donor to the electron transport chain, how many moles of FADH2 would be required to produce a proton gradient in which exactly one mole of protons have been pumped across the membrane, assuming we start with no gradient? The standard reduction potential of FADH2 is +0.10 V, and that of O2 is +0.81 V. Select the closest value from the options below. A) 3 mol FADH2 C) 0.17 mol FADH2 B) 1 mol FADH2 D) 5.8 mol FADH21. As has been discussed in class the electron transport chain is the first part of oxidative phosphorylation. This chain of multi-subunit enzymes is responsible for facilitating the transfer of electrons from NADH and succinate/FADH₂ to oxygen making water. Given this understanding answer the following questions. a. Below is an equation for determining the amount of energy required to move 1 mol of protons across the inner membrane of the mitochondria. In actively respiring mitochondria the A = +/-0.15 volts to +/- 0.20 volts. The ApH for a typically mitochondria will be 0.75 pH units, with the matrix (N-side) being more alkaline than the IMS (P-side). Given this state the total energy needed to move a single proton from the N to the P-side. Assume T = 310K and R = 8.314 J mol-¹ K-¹ and a A4 = 0.17 volts. Finally, assume F = 96.5 kJ/mol/volt AG = 2.3RTAPH + FA b. How much free energy is available in the transfer of electrons from NADH to oxygen given the following half-reactions and…1a. Name and draw diagrammatically the series of mitochondrial electron transfer catalysts, starting with the oxidation of NADH and succinate and ending with the reduction of O2. Indicate the sites and stoichiometry (per 2e) at which protons are translocated from the matrix to the intermembrane space. c. Indicate which complexes are inhibited by: amytal, antimycin A, azide (N3), cyanide (CN), carbon monoxide (CO), and rotenone. d. Describe the effects of (1) oligomycin and (2) uncouplers of oxidative phosphorylation, e.g., dinitrophenol (DNP), carbonyl cyanide-p- trifluoromethoxyphenylhydrazone (FCCP), on respiration and ATP synthesis when added to a suspension of mitochondria with excess malate, ADP, and inorganic phosphate (Pi). b. I
- 1. Name and draw diagrammatically the series of mitochondrial electron transfer catalysts, starting with the oxidation of NADH and succinate and ending with the reduction of O2. 2. Indicate the sites and stoichiometry (per 2e) at which protons are translocated from the matrix to the intermembrane space. 3. Indicate which complexes are inhibited by: amytal, antimycin A, azide (N3 ), cyanide (CN), carbon monoxide (CO), and rotenone. 4. Describe the effects of (1) oligomycin and (2) uncouplers of oxidative phosphorylation, e.g., dinitrophenol (DNP), carbonyl cyanide-p- trifluoromethoxyphenylhydrazone (FCCP), on respiration and ATP synthesis when added to a suspension of mitochondria with excess malate, ADP, and inorganic phosphate (Pi).In the 1930s, some physicians prescribed low doses of a compound called dinitrophenol (DNP) to help patients lose weight. This unsafe method was abandoned after some patients died. DNP uncouples the chemiosmotic machinery by making the lipid bilayer of the inner mitochondrial membrane leaky to H+. Chemical agents that cause this effect are called uncouplers. Explain how this could cause weight loss and also death. Considering the danger, is there any use for compounds like DNP or other uncouplers?9. Two nutrient solutions are maintained at the same pH. Actively respiring mitochondria are isolated and placed into each of the two solutions. Oxygen gas is bubbled into one solution. The other solution is depleted of available oxygen. Which of the following best explains why ATP production is greater in the tube with oxygen than in the tube without oxygen? (A) The rate of proton pumping across the inner mitochondrial membrane is lower in the sample without oxygen. (B) Electron transport is reduced in the absence of a plasma membrane. (C) In the absence of oxygen, oxidative phosphorylation produces more ATP than does fermentation. (D) In the presence of oxygen, glycolysis produces more ATP than in the absence of oxygen.
- 15. In the ETC shown below, explain, IN YOUR OWN WORDS, what is occurring at the different stages labeled. a. b. C. d. e. Electron transport chain ATP synthase Intermembrane space Inner mitochondrial- membrane NADH A (H+ NAD + H+ Mitochondrial matrix B H+ H+ (H+ FADH₂ FAD+ H+ (H+ (H+ 2 free 2 electrons hydrogen exiting ETC ions C H+ H+ 1/2 of an O₂ molecule (H+ H+ (H+ (H+ H₂O x 2 H ADP PO (H (H+ D E ATP2. The diagram on the right taken from the textbook il- lustrates the structure of a mitochondrion in a mamma- mitochondrial H* H+ lian cell containing the enzymes responsible for catalyz- membrane. ing the oxidation of metabolites coupled to the reduction of O2 and the synthesis of ATP. Only the inner mito- chondrial membrane is shown to simply the dia- gram and discussion. Sonication produces submito- chondrial particles in which the outer membrane has been stripped away and the inner membrane forms a closed vesicle known as an "inside-out particle". That is, the intermembrane space becomes the lumen of the particle, and the membrane bound components that faced the matrix in the intact mitochondrion are now ex- Electron carriers (respiratory chain) Inner H+ H+ H+ H+ H+ + H+ H+ 02 (e- acceptor) 3 Energy of e- flow stored as electrochemical potential. Reduced e- donor H+ ATP ADP + P, H+ synthase H+ H+ АТР H+ + H+ H+ H+ H+ H+ H+ H+ H+ H+ posed to the solvent in which the particles are…3. Draw the structures of glucose, pyruvate, and acetyl coenzyme A; number (label) the carbons in each. Glucose Pyruvate Acetyl-CoA 4. Assuming a typical mitochondrial matrix has a volume of about 5x10-¹4mL. If the pH of the matrix is 7, report the number of hydrogen ions contained in the matrix. ANSWER 5. Glucose may be used to produce pyruvate or stored. Under what circumstances would glucose be stored?