LECTURE 23 - MIDTERM 3 Flashcards
What is oxidative phosphorylation?
– it is the coupling of energy release during electron transport to ATP synthesis
– mechanism of oxidative phosphorylation is converting free energy of proton gradient for ATP synthesis
Describe the Chemiosmotic Theory and it’s components.
– now widely accepted model for mechanism of oxidative phosphorylation
– states that free energy from electron transport drives an active transport system which pumps protons out of mitochondrial matrix into intermembrane spcae
– this action generates an electrochemical gradient for protons
– protons on the outside have thermodynamic tendency to flow back in down electrochemical gradient and this flow provides driving force for ATP synthesis
What is ATP synthase?
– a reversible coupling device that can convert the energy of the electrochemical proton gradient into chemical bond energy or vice versa
- the direction depends on the magnitude of the electrochemical proton gradient
What is the binding-change mechanism for ATP synthesis by ATP synthase?
– the beta subunit exists in three conformation states:
L- losse - binds ADP and Pi “loosely”
T - tight - catalyzes formation of ATP from bound ADP and Pi – the newly made ATP binds with very high affinity and is not released until the Beta subunit assumes the O conformation
O - open - ATP can now be released and ADP and Pi can enter the enzyme
– the conformation state of the beta subunit (L, T, or O) is determined by contact with the gamma subunit
What is the conformation state of the beta subunit based on?
– dtermined by contact with the gamma subunit
– passage of protons through channels in F0 causes gamma to rotate counter-clockwise
– rotation of the gamma subunit allows a given beta subunit to change conformational states
– rotation is counter-clockwise
T or F, rotation of the gamma subunit allows cycles of ATP synthesis and release through changes to the Beta subunit conformation
True;
subunit 1: L –> T –> O –> L –> T –> O
subunit 2: O–> L–> T –> O –> L –> T
subunit 3: T –> O –> L –> T –> O –> L
What is rotating in F1?
– gamma is rotating in F1, everything else stays the same
How does the proton-motive force drive rotation of the F0 assembly?
– an aspartic acid residue in the c subunit and two half-channels in the “a” subunit are key
What is a key component of half-channels on the ATP synthase?
– channels do not span the membrane
– channels have an aqueous hydrophilic character (thus protons can flow into/out of the channels)
– aspartic acid positioned so that it comes into contact with two different environments –> interacts w/ two different environments
1: the hydrophilic half-channels in subunit a
2: hydrophobic environment of the membrane
T or F, proton is 1 to 1 for C subunits w/ aspartic acid
– True, 1 aspartic acid; 1 proton
What is a key feature of the aspartic acid in the half channel?
– the charge on the aspartic acid changes depending on the environment
– in a hydrophilic environment, the residue can be in the aspartate form (deprotonated) –> base form
– in the membrane, the residue is protonated and can exist in a hydrophobic environment –> acid form
Describe the rotation of the C subunit.
– the intermembrane space half-channel is proton-rich, and thus the aspartic acid becomes protonated
– protonation of the C subunit causes the entire ring to turn to moving Asp to the hydrophobic membrane
– this positions a different c subunit next to the matrix half-channel, which is proton-poor and thus the aspartic acid is deprotonated
– then the cycle repeats
What is the key point of the rotation of the C subunit?
– key point is that the proton gradient ensures protonation of the aspartic acid in the intermembrane half-channel
– causing movement
– and deprotonation in the matrix half-channel
– which resets the system
T or F, the C subunit rotation clockwise means it’s hydrolyzing ATP and counterclockwise means it’s synthesizing ATP
False;
– clockwise = synthesizing ATP
– counter clockwise = hydrolyzing ATP
T or F, there’s one matrix half-channel and one cytoplasmic half channel per ATP synthase
True
How do protons enter the ATP synthase?
– protons enter from the intermembrane side, then ride around, and are released on the matrix side
– proton binding causes rotation, proton release resets the system to allow additional binding and additional rotation
– protons that are released in the matrix get pumped out again
What allows the release of ATP from the ATP synthase?
– rotation of the c ring turns the gamma subunit, which causes conformation changes to the beta subunits, which allows release of ATP
What is the point of oxidative phosphorylation?
– to generate ATP
How is ATP transported out of the matrix? And how is ADP brought in?
– ATP-ADP translocase (specific transport protein) allows a 1:1 exchange of ADP for ATP
– ATP-ADP translocase is in high abundance and it makes up about 15% of the protein content in the inner mitochondrial membrane
T or F, ATP transport is energetically expensive due to its negative charge
True; ATP has one more negative charge than ADP – this dampens the membrane potential
– difference in charges needs to be maintained for proton gradient
Where does 25% of the energy yield from respiration go?
– goes back into regenerating the membrane potential
– part of the proton pumping we did during electron transport is used to maintain the electrical gradient
– therefore, some protons aren’t used to make ATP directly
What is the importance of mitochondrial shuttles?
– mitochondrial shuttles allow transport across inner membrane
– inner membrane is extremely impermeable yet exchange w/ the cytoplasm is important for respiration
– solution = mitochondrial shuttles
T or F, functionally, transport of OH- out is the same as transport of H+ in
True
T or F, NAD+ in the cytoplasm must be regenerated for use during glycolysis (cytoplasm)
True
T or F, ETC is happening in the inner membrane of the mitochondria
True
T or F, the glycerol 3-phosphate shuttle transfers electrons (but not NADH itself) across the inner membrane
True; electrons are transferred to FAD and NADH is oxidized back to NAD+ without leaving the cytoplasm
What is the whole point of the glycerol 3- phosphate shuttle? And what are the steps that occur?
– whole point of shuttle is to take NADH from cytoplasm for it to be used in ETC
glycerol 3-phosphate is located on inner mito membrane (is associated with FAD in a similar way to complex 2)
- Reduction of DHAP by NADH in cytosol
- Reoxidation of G3P and reduction of FAD
- Transfer of an electron pair from FADH2 to coenzyme Q (in complex III)
- DHAP returns to the cytosol
What is the malate-aspartate shuttle?
– a more complicated shuttle that regenerates cytoplasmic NAD+ in heart, kidney and liver tissue
What are the steps that occur in the Malate-Aspartate shuttle?
– Malate Dehydrogenase reduces Oxaloacetate to Malate and accepts e- from NADH
– Malate shuttle brings Malate across inner membrane
– Malate is reoxidized by Malate dehydrogenase and transfers e- back to NAD+ on Matrix side
– Oxaloacetate is transaminated to make Aspartate, which can transverse back across the membrane using the Aspartate shuttle
– Aspartate is converted back to Oxaloacetate
