ATP synthase Flashcards
Whats the difference between white and dark mest
NUmber of mitochondria per cell will depend on the energy requirements of the cell. Density in mitochondria muscles contribute to the difference in coloratioin of meat.
How do protons get back into the matrix
- Through ATP synthase
High membrnae potential -> high ATP synthesis
2. Through uncoupers (heat-generated as side-effect)
Lower membrane potential -> reduced ATP synthesis
What are the methods to elucidate a protein structure
- x-ray crystallography
forms form crystal + x-ray - Cryo-electron microsocpy
freeze protein -> beam of electrons is fired -> take photos ->
where is the mushroom head of ATP synthase
F1
What are the 2 motors of ATP synthase
F0: electric motor (transmembrane)
F1: chemical motor (matrix)
Where is the F0 subunit located? What subunits are there in F0?
F0 subunit is embedded in the inner mitochondrial membrane
Made by subunits a,b and c
Where is the F1 subunit located? What subunits
F1 subunit contains the catalytic activity of the synthase
Made by 3 alpha, 3 beta, 1 gamma, 1 epsilon, 1 delta
Beta: active site, each subunit has different interactions with the gamma subunit
gamma and epsilon form the stalk
Whats the structure of the proton channel F0? Amino acids? how does it connect to F1
Ring composed of 12 identential c chains (number can depend on the speicdes)
Asp or Glu is found in the middle of the helices (negatively charged)
Arg is found in the a sub unit (positively charged)
Connected to F1 through central gamma and epsilon and exterior column (b in orange)
What is the structure of the catalytic unit F1
5 subunits: 3 alphas, 3 betas, 1 gamma, 1 epsilon, 1 delta.
Uses the power of rotational motion to build ATP
3 steps are needed:
1. Trapping of ADP and Pi
2. Formation of new phosphate bond
3. Release of ATP and recharge
ATP synthase tends to form dimers?
Mitochondrial ATP synthase forms dimers. Asosciation stabilizes individual enzymes to the rotational forces required for catalysis and promotes curvature of memrbane
F0 subunit rotates in which direction as viewed from the top
Clockwise (intermembrane space)
explain the motive force for ATP synthase
ATP synthase uses a proton motive force as a source of energy => drives mechanical rotatry mechanism that leads to the chemical synthesis of ATP from ADP and Pi
F1 direction as viewed from the matrix
Counterclockwise
What are the important amino acids in the c subunit that are essential for protein translocation?
Ionizable glutamic acid
aspartic acid
and the arginin in the a subunit -> gives initial push
What is the protononation of glutamate, aspartate
Glutamate <-> glutamic acid
Aspartate <-> aspartic acid
How do protons flow across the inner mitochondrial membrane
When a proton hops on, the Glu or Asp residue in the c subunit is neutralized and the ring can step forward.
The arrival of one proteon through one half channel is paralleled by the exit of another through the half channel
Subunit a, which is next to the c subunit, has 2 channels that reach halfway into the a subunit. One half channel opens to the intermembrane space and the other to the matrix.
The movement of protons through the half channels from the high proton concentration of the inner membrnae space ot the low proton concetntration of the matrix powers the rotation of the c ring
Glutamate or aspartate is found in the middle of the c subunit.
If glutamate is charged (unprotonated) -> c subunit does not move into hydrophobic interior of membrnae
If glutamate is charged (unprotonated)
there is no rotation
Explain step by step how protein flow drives rotation of c ring
Protons enter the half channnel of the subunit a from the positive side of the membrane
Proton that entered subunit a is loaded into 1 c subunit
Proton binds to a glutamate or aspartate residue on one of the subunits of the c ring
-> allow it to rotate clockwise
glutamic or aspartic acid in the in the other half channel releases protons inside the matrix
Rotation bring s the deprotonated subunit to the postion where it can eb loaded with another proton from the intermembrane space
How is the rotation of the F0 subunit connected to the F1 subunit
As the c ring rotates, the gamma and epsilon subunits turn inside the hexamer of the F1 subunits.
Rotation of the gamma subunit promotes syntehsis of ATP
The exterior column formed by the b chains and the delta subunit anchor the hexamor and prevent it from movign
How does the gamma subunit interact with teh beta subunit
gamma subunit is the knob that rotate s-> squeezing the beta subunits
What are the alternate configurations of the beta subunits
Lose -> tight -> open
What mechanism accounts for the syntheiss of ATP
Binding change mechanism
the 3 catalytic beta subunits of the F1 component can exist in three conformations
Lose form: nucleotides are trapped in the beta subunit. Binds ADP + Pi
Tight form: ATP is synthesized from ADP and Pi
Open form: nucleotides can bind to or be released from the beta subunit
What rotation can drive the interconversion between the 3 forms? How many grees
Rotation of the gamma subunit drives the interconversion of the 3 forms by altering their substrate binding abilities
explain what happens in the L, T, O configurations
Loose: ADP and Pi in the L form trap the substrates
Tight: ADP and P formed in the T-form combine to form ATP
Open: ATP in the O site can now depart of the enzyme to be replaced by ADP and pi
Summarize the F1 subunit
No 2 subunits are ever in the same confomration
Each subunit cycles through the three conformations
Hexamer does not move, gamma rotates in 120 increments. Each 120 rotation releases 1 ATP
1 full rotation -> 3 ATPs are rleased
Yest: 10 c subunits
chlorpoast: 14 subinits.
Which is more effective
yeast is more effective.
More c subunits -> more protons needed for a full rotation
what needs to be taken into account when calculating the ATP yield
Where the metabolites are made
Number of protons translocated into intermembrane space by ETC
Number of protons required to make F0 complete 1 full rotation
Number of ATPs produced by F1 after 1 full rotation
What does ATP synthase need to make ATP
- Protons from NADH and FADH2 pumped into intermembrane space (inside mitochondria)
- ADP in the matrix
- Phosphate in the matrix
What shuttles are used to move cytoplasmic NADH
A. Glycerol-3-phosphate shuttle
B. Malate-aspartate shuttle
To move ADP, what carrier is used?
ADP translocase
To move Pi
Phosphate carrier
How does glycerol 3-phosphate shuttle work?
Which tissue
In the muscle and brain
Electrons from cytoplasmic NADH can enter the ETC by using glycerol phosphate shuttle.
DHAP <-> glycerol -3 phosphate (via cytoplasmic glycerol -3 -phosphate dehydrogenase)
in the process NADH H -> NAD+ and the electron is transfered to glyerol 3 phosphate.
glycerol 3 - p <-> DHAP (via mitochondrial glycerol-3-phosphate dehydrogenase)
In the process E-FAD is convereted to FADH2
Later electron is passed to Q <-> QH2
This leads to a loss of energy, since only 6H+ are pumped
How many protons are pumped from NADH in the cytosol
6H+ per NADH cytosol
Do not get confused between?
Glycerol -3 phosphate
Glyceradehyde -3-phosphate
Can NADH be carried across the membrnae?
No: instead of carrying NADH across membranes, transfer electrons from NADH through shuttles instead transfer electrons from NADH through shuttes
how does the malate-aspartate shuttle work?
The malata asparatate shuttle consist of 2 membrnae antiporters, 4 enzymes including 2 malate dehydrogenases.
Glutamatate, Aspartate are moved in/out
Malate/alphaketoglutarate moves in out
Malate moves in (NADH -> NAD+, hydrogen is on malate)
once in, malate -> OAA (NAD+ <-> NADH)
alpha ketoglutate moves out
Which tisuses uses the malate-aspartate shuttle
Heart, kidney and liver
How much H+ per NADH from cytosol
10 H+
How is ADP entry coupled to ATP exit by ATP-ADP translocase?
The translocase catalyzes the coupled entry of ADP into thematrix and the exit of ATP from it.
Binding of ADP from the cytoplasm favors the eversion of transproters to release ADP into the matrix
Subsequent binding of ATP from the matrix to the everted form -> favors eversion back to the original form -> releasing ATP into cytoplasm
For each ATP synthesized, a phosphate is required inside the matrix. How does the phosphate get in? whats the cost
Each phosphate inside the matrix, requires an additional H+ inside the matrix, reducing the proton gradietn.
It adds 1 H+ to the cost of 1 ATP
Whats the final calculation? How mnay H for 3 ATP
How many H for 1 ATP
How many ATPs per NADH
How many ATPs per FADH2
13 H+ for 3 ATPs
4H+ for 1 ATP
10/4 ATPs per NADH
6/4 = 1.5 ATPs per FADH2
Recap the amount of NADH/FADH2 made per glucose
Glycolysis: 2 NADH
PDH: 2 NADH
Citric acid cycle: 6 6NADH, 2 FADh2
Electron transport chain: 1 NADH = 10H+
1 FADH2 = 6H+
Whats the energy yield from oxidative metabolism (in muscle cells)
+30
How do NADH/FADH2 ~ ATP regulate each other?
NADH and FADH2 are oxidized when ATP is made
The synthesis of ATP from ADP and Pi control the flow of electrons from NADH and FADH2 to oxygen
The availbaility of NAD+ and FAD control the rate of the citric acid acid
When ADP concnetration rises, the rate of oxidative phosphorylation increases to meet the ATP needs in the cell.
What are the roles of the mitochondria
- Aerobic energy production
- Reactive oxygen species formation
What are some neurological disorders implicated with defects in mitochondrial dysfunction
Alzeihmers, Autism, epilepsy, lou hehirdge
what ETC complexes have reduced activity in autistic individuals
Complex I and 5
What does Mt dysfunction do?
Mt disfunction results in less energy available to fuel the high energy needs ot the brain and muscles, and leaves free radicals in the system where they can cause damage.
Not all autism Spectrum Disorders involve mitochondrial disease.
Potential targets for the association between Autism Spectrum Disorders and Mt dysfunction?
Changes in diet or supplements
Fat accumulation under different temperatures. PET-CT scan shows uptake and distribution of FDG under different temperatures. What is different?
Femailes have hgiher accumulation of borwn adipose tissue -> higher mitochondrial activity, use the proton gradient to generate heat
Difference between white fat and brown fat
White fat: stores energy in large lipid droplets, provides insulation
Brown fat: Rich in UCP1. Helps maintain body temperature by thermogensis
How do uncoupling proteins and thermogenesis work?
Signalling cascade leads to release of fatty acids from lipid droplets.
Fatty acids activate UCP1 in mitochondria
UCP1 allows the leak of protons dissipating the protein gradient
Energy from oxidation of fuels is released as heat
What happens during a grizzly bears winter sleep
- Body temperature drops slightly
- Metabolic acitivty is very low
- do not eat or drink
- they lose body fat every day
How does thermogenesis work in grizzly bears?
UCP1 is used by animals to control thermogenesis
Cell-specific expression of the UCP1 protein leads to heat production under aerobic conditions.
During the final weeks of hibernation, norepinephrine (hormone) level increaes and activates UCP1 expression at the base of the neck -> warms up the blood travelling to the brain andd signals the end of a winter nap.
UCP1 allows protons to cross the inner mt membrane and thereby uncouple electron transport from ATP synthesis.
