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In the preparation of inside-out particles, the F1 component of the ATP synthase enzyme (a) can pe reversibly removed. Suppose that the F1 component has been removed and the pH of the external medium was made equal to that of the matrix prior to sonication. What will be the pH of the lumen of re-suspended particles devoid of the F1 component? (b) wish to test for the intactness of the electron transport chain. Which electron transfer catalysts may Because the suspension of isolated inside-out particles is exposed to the atmosphere, you have become diluted in the preparation protocol and have to be replenished in the suspension to achieve observable rates of Oz reduction? (c) electron transfer chain can be added to test for intactness of the ETC by reduction of O2? Explain briefly the basis of your answer. Having added the electron transfer catalysts in part (b) above, which two substrates of the (d) out particles. Using either of the substrates in your answer to part (c) above in addition to ADP + Pi added to the suspension, will ATP synthesis increase or decrease with an increase in pH of the solvent in which the particles are suspended? Explain. (e) mislabeled solutions and you do not know which of the two substrates mentioned in part (c) was added. What would be the most straightforward way to identify which substrate was added to observe reduction of O2 coupled to ATP synthesis? The F1 component of the ATP synthase enzyme is now added to the suspension of inside- Your lab technician was requested to complete the series of experiments; however, he/she

In the preparation of inside-out particles, the F1 component of the ATP synthase enzyme (a) can pe reversibly removed. Suppose that the F1 component has been removed and the pH of the external medium was made equal to that of the matrix prior to sonication. What will be the pH of the lumen of re-suspended particles devoid of the F1 component? (b) wish to test for the intactness of the electron transport chain. Which electron transfer catalysts may Because the suspension of isolated inside-out particles is exposed to the atmosphere, you have become diluted in the preparation protocol and have to be replenished in the suspension to achieve observable rates of Oz reduction? (c) electron transfer chain can be added to test for intactness of the ETC by reduction of O2? Explain briefly the basis of your answer. Having added the electron transfer catalysts in part (b) above, which two substrates of the (d) out particles. Using either of the substrates in your answer to part (c) above in addition to ADP + Pi added to the suspension, will ATP synthesis increase or decrease with an increase in pH of the solvent in which the particles are suspended? Explain. (e) mislabeled solutions and you do not know which of the two substrates mentioned in part (c) was added. What would be the most straightforward way to identify which substrate was added to observe reduction of O2 coupled to ATP synthesis? The F1 component of the ATP synthase enzyme is now added to the suspension of inside- Your lab technician was requested to complete the series of experiments; however, he/she

Biochemistry
Biochemistry
9th Edition
ISBN:9781319114671
Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.
Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.
Chapter1: Biochemistry: An Evolving Science
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2. 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 sus- pended. Differential centrifugation allows separation of the inside-out particles from the outer mem- brane fragments and soluble TCA enzymes. Assume for this question that sonication does not change the catalytic H+ H+ H+ H+ erties of any membrane bound components. In the preparation of inside-out particles, the F1 component of the ATP synthase enzyme (a) can pe reversibly removed. Suppose that the F1 component has been removed and the pH of the external medium was made equal to that of the matrix prior to sonication. What will be the pH of the lumen of re-suspended particles devoid of the F1 component? Because the suspension of isolated inside-out particles is exposed to the atmosphere, you (b) wish to test for the intactness of the electron transport chain. Which electron transfer catalysts may have become diluted in the preparation protocol and have to be replenished in the suspension to achieve observable rates of O2 reduction? (c) electron transfer chain can be added to test for intactness of the ETC by reduction of O2? Explain briefly the basis of your answer. Having added the electron transfer catalysts in part (b) above, which two substrates of the (d) out particles. Using either of the substrates in your answer to part (c) above in addition to ADP + Pi added to the suspension, will ATP synthesis increase or decrease with an increase in pH of the solvent in which the particles are suspended? Explain. The F1 component of the ATP synthase enzyme is now added to the suspension of inside- (e) mislabeled solutions and you do not know which of the two substrates mentioned in part (c) was added. What would be the most straightforward way to identify which substrate was added to observe reduction of O2 coupled to ATP synthesis? Your lab technician was requested to complete the series of experiments; however, he/she
Transcribed Image Text:2. 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 sus- pended. Differential centrifugation allows separation of the inside-out particles from the outer mem- brane fragments and soluble TCA enzymes. Assume for this question that sonication does not change the catalytic H+ H+ H+ H+ erties of any membrane bound components. In the preparation of inside-out particles, the F1 component of the ATP synthase enzyme (a) can pe reversibly removed. Suppose that the F1 component has been removed and the pH of the external medium was made equal to that of the matrix prior to sonication. What will be the pH of the lumen of re-suspended particles devoid of the F1 component? Because the suspension of isolated inside-out particles is exposed to the atmosphere, you (b) wish to test for the intactness of the electron transport chain. Which electron transfer catalysts may have become diluted in the preparation protocol and have to be replenished in the suspension to achieve observable rates of O2 reduction? (c) electron transfer chain can be added to test for intactness of the ETC by reduction of O2? Explain briefly the basis of your answer. Having added the electron transfer catalysts in part (b) above, which two substrates of the (d) out particles. Using either of the substrates in your answer to part (c) above in addition to ADP + Pi added to the suspension, will ATP synthesis increase or decrease with an increase in pH of the solvent in which the particles are suspended? Explain. The F1 component of the ATP synthase enzyme is now added to the suspension of inside- (e) mislabeled solutions and you do not know which of the two substrates mentioned in part (c) was added. What would be the most straightforward way to identify which substrate was added to observe reduction of O2 coupled to ATP synthesis? Your lab technician was requested to complete the series of experiments; however, he/she
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