OxPhos

Question 1

DeltaG =

Answer 1

-nFE

• n = number of electrons transferred
• F = Farradays Constant = 23 kcal/Volt
• E = change in reduction potential

Question 2

What does it mean if something. Has a negative. Reduction potential? Positive?

Answer 2

If it is negative then it will be likely to give up electrons
If it is positive then it wants to take up electrons.

Question 3

The total free energy made in cellular respiration is 53 kcal/mol but we only get 3 ATPs out, which each cost 7.3 kcal to make. Where does the rest of it go?

Answer 3

it is lost as heat and is used to maintain body temp.

Question 4

Complex 1

Answer 4

• Also called NADH Dehydrogenase
• It accepts electrons from NADH in the mitochondrial matrix and pumps protons out into the inter membrane space.
• Through this, NADH gets oxidized to NAD+ so that it is ready for another round of the TCA cycle (Complex 1 gets reduced)
• Has a Flavin Mononucleotide (FMN) and many Iron-Sulfer centers
• Complex 1 will then transfer those electrons over to CoQ10

Question 5

Complex 2

Answer 5

• Also called Succinate Dehydrogenase
• Complex 2 is reduced by FADH2 from the TCA cycle.
• Complex 2 then shuttles the electrons over to CoQ10
• There are no protons pumped into the inter membrane space this time.

Question 6

Bypass Reactions

Answer 6

CoQ can also accept electrons from the FADH2 via

• Glycerol-3-phosphate
• fatty acyl CoA dehydrogenase

Question 7

Where is complex 2 located? How about glycerol-3-phosphate Dehydrogenase? Fatty acyl CoA dehydrogenase?

Answer 7

Complex 2 - matrix side of the inner membrane
Glycerol-3-phosphate dehydrogenase - inter membrane side of the inner membrane
Fatty acyl CoA dehydrogenase - In the matrix

Question 8

Complex 3

Answer 8

• Receives electrons from CoQ (Complex 3 is reduced)
• Passes those electrons over to cytochrome-c
• consists of cytochrome-b, cytochrome-c1, and an Fe-S center.
• Protons are pumped into intermembrane space in this step.

Question 9

What is the result of the Lowering of pH in the intermembrane space?

Answer 9

There is a proton-motive force generated, which activated the Fo part of the ATP synthase (complex 5). This will begin the process of making ATP from ADP.

Question 10

What are the prerequisites for oxidative phosphorylation to occur??

Answer 10

1) Availability of NADH and FADH2 as reducing agents.
2) pH difference
3) Presence of oxygen as a terminal oxidizing agent
4) High ADP/ATP ratio
5) sufficient quantity of mitochondria with proper enzymes

Question 11

How does the ADP get into the mitochondrial matrix for it to eventually get turned into ATP?

Answer 11

There is an ATP/ADP antiporter, which also requires the pH gradient and the membrane potential in order for it to be able to shuttle the ADP in the mitochondrial matrix.

Question 12

Complex 4

Answer 12

• Also known as cytochrome oxidase
• final component of the ETC
• It accepts electrons from cytochrome c
• coordinates the transfer of 4 electrons to oxygen, reducing it to water
• contains cyt a, cyt a3, and 2 copper ions.

Question 13

What would happen in a low oxygen situation?

Answer 13

Complex 4 wouldn’t be able to shuttle its electrons over to oxygen to make H20 –> Therefore, ATP will not be made –> Therefore Na/K ATPases won’t be made –> Therefore Na will stay in cells –> Also, the H+ gradient will just go down –> Cells will die.

All of these things will increase the calcium concentration, which increases mitochondrial permeability.

Question 14

Atractyloside

Answer 14

Disrupts the ADP/ATP antiporter, so no ADP will be brought into cell. Therefore, ATP synthesis will be disturbed.
- happens often in the Middle East and in Africa because there is a flower that looks like an artichoke.

Question 15

Amytal

Answer 15

• reversible inhibitor of Complex 1
• also known as truth serum, it is a barbiturate.
• used to treat anxiety, insomnia, and epilepsy.
• Has been shown to protect cardiac muscle during ischemia-repercussion by limiting the production of ROS
• In low concentrations it is OK, in high concentrations it is lethal.

Question 16

Rotenone

Answer 16

• Also inhibits complex 1

- also known as fish poison.

Question 17

Why is it that inhibiting complex 1 or 2 you might still be able to get some ATP out?

Answer 17

Because if you remember, Complex 1 and 2 are independent of each other. On the contrary, everything else work in concert so if you block 3,4,or 5 there will be no ATP.

Question 18

Antimycin

Answer 18

• Inhibitor of Complex III
• anti-fungal used in agriculture
• no ATP will be made
• binds to cytochrome b in the reduced state.
• – if you think about it, normally every step in the process gives over its electrons to the next so they all end up oxidized. Here though, there is backlog, so Complex 1,2, and CoQ will be reduced and Cyt c and Complex 4 and oxidized.

Question 19

Cyanide (CN) poisoning

Answer 19

Cyanide will bind to the heme iron in its oxidized form (Ferric iron - Fe3+) in the cyt a3 component of complex 4 and not let it get reduced. Therefore, cyt c will just be holding on to the electrons and not be able to give them over.

• all ETC is stopped and no ATP is made
• Complex 1,2, CoQ, 3, Cyt c will all be in reduced form

Question 20

How to treat CN poisoning

Answer 20

Nitrate followed by thiosulfate treatment.
- the reason that this works is because the nitrate will convert Hb into MetHb, which has a higher affinity for CN. Then, we give the patient thiosulfate, which will cause conversion into thiocyanate through the enzyme Rhodanese. Thiocyanate can then be excreted from the kidneys.

Question 21

Oligomycin

Answer 21

• Binds to the Fo portion of ATP synthase, which effectively blocks the reentrant of protons into the matrix of the mitochondria.
• No ATP is made and the proton gradient builds up.
• electron transport will stop too because of the difficulty in pumping protons against the steep proton gradient.

Hint: The “o” in Fo stands for Oligomycin

Question 22

Uncouplers

Answer 22

• molecules that allow protons to renter the mitochondrial matrix from the intermembrane space
• phosphorylation of ADP becomes uncoupled from the transfer of electrons.
• respiration rate will be every high because of ADP/ATP ratio.
• the rate of respiration is limited only by availability of NADH, Succinate and O2.
• You will see acceleration of both the TCA cycle and electron transfer to O2.
• Inhibition of ATP synthase (because no gradient)
• generation of heat

Question 23

What are some examples of Uncouplers?

Answer 23

AraC, AZT - both of these damage the membrane and allow protons to flow across.
DNP, high dose aspirin - lipid soluble proton carriers than can pick up protons from the intermembrane space and transport them across to the matrix.
Proton channels

Question 24

What re two ways we generate heat?

Answer 24

Shivering and BAT

Question 25

When you begin to. Feel Cold what is the mechanism that. Heats you up (not shivering)

Answer 25

1) Norepinephrine is released from sympathetic nerve endings. 2) Norepi activates a lipase in BAT that cleaves TAGs into fatty acids. 3) Fatty acids activate UCP-1 while also serving as fuel to make ATP. 4) UCP-1 acts to allow protons to come back across the inner membrane, thus causing no ATP generation but heat production will occur.

Question 26

Malate-aspartate shuttle

Answer 26

Operates in the heart, kidney, and liver - Generates NADH in the mitochondrial matrix, which can then enter complex 1. - this only works if the NADH/NAD+ ratio is higher in the cytosol than the matrix. - Here, 1 NADH = 3 ATP Mechanism: Oxaloacetate gets broken down into Malate. Malate then enters the mitochondrial matrix via a Malate/alpha-ketoglutrate anti-port. Malate then donates its electrons to NAD+ to become NADH and in the process becomes Oxaloacetate. Oxaloacetate then meets up with glutamate to become alpha ketoglutarate and aspartate. The alpha ketoglutarate will go back out to the cytosol via the cytosol that we talked about. The aspartate will go out to the cytosol via another anti port between aspartate and glutamate

Question 27

Glycerophosphate shuttle

Answer 27

- operates in skeletal muscle and brain - FAHD2 in the inner mitochondrial membrane donates its electrons to CoQ in the inner mitochondrial membrane. - Here, one NADH = 2 ATP Mechanism: Basically Glucose will get broken down DHAP, which will be converted into Glycerol-3-Phosphate. It will enter the outer leaflet of the mitochondrial matrix. Then it meets up with Glycerol phosphate shuttle, which will take the electrons and give them to FADH2, converting G3P into DHAP. FADH2 will then use them as a bypass rxn in the ETC. DHAP will. Come back out into the cytosol.

Question 28

Inner membrane vs outer membrane

Answer 28

Inner - impermeable to small molecules (except O2, CO2, H20, NH3) and has transporters in it. - Has the ETC on it Outer - permeable to small molecules and actually has poring for them to get through.