Unit 8 - Oxidative Phosphorylation

Question 1

What is oxidative phosphorylation?

Answer 1

The formation of ATP as a result of the transfer of eldctrons from NADH and FADH₂ to O₂ by carrier in the ETC

Question 2

What is the core idea of oxidative phosphorylation?

Answer 2

Create a proton gradient using the high electron transfer potential of the NADH and FADH₂ carriers to create a electrochemical gradient from the IMM into the mitochondrial matrix. This create a Proton moving force that drives ATP synthase to create ATP

Question 3

What are the 3 ways in which electrons can be transfered as?

Answer 3

Free electrons
hydrogen atoms

H- (hydride ions)

Question 4

How can we measure reduction potential?

Answer 4

Using standard reduction potentials - the more negative, the more willing something is to reduce/accept electrons

Question 5

How many ATP molecules can we produced from NADH in theory? In practice?

Answer 5

In theory, 7

In practice, 2.5 ATP/per NADH

Question 6

What is complex 1 of the ETC, and what does it do?

Answer 6

NADH-Q Oxidreductase

NADH is reduced, giving 2e- (accepted by FMN in oxidreductase)

Electrons transfered to FE-S clusters, and finallly to co-enzyme Q (Which is reduced); i.e. ubiquinone to ubiquinole

This pumps 4 protons into IMS from matrix

Question 7

Net equation of Complex 1?

Answer 7

Question 8

What is complex 2 and what does it do?

Answer 8

Succinate-Q oxidreductase (AKA succinate Q reductase)

Succinate dehydrogenase, from Kreb cycle, is part of this complex

Succinate is Oxidized (hence it’s first)

electrons transfered via FADH₂ within the complex

Q reduced to QH2 (ubiquintine to ubioquinol via FeS clusters)

NO protons are pumped

Question 9

What is coenzyme Q? (i.e. structure)

What does it do?

Answer 9

Small hydrophobic molecule in IMM, contains repeated isoprenoid tail (type of hydrocarbon)

Accepts 2e- to reduce to ubiquinole (from ubiquinone)

Acts as electron shuttle for electrons from complex I and II & carries to complex III

Question 10

What is complex III and what does it do?

Answer 10

Q-cytochrome C oxidreductase

Takes 2 electrons from QH₂ (oxidation) and transfers it individually to FeS clusters to heme C₁ then to cytochrome c (reduction)

Pumps 4 protons into IMS

2 come from QH₂ (Which came from the matrix going further back)

2 come from matrix

Question 11

What are the 2 key cytochromes in complex III?

Answer 11

cyt b

cyt c1

Question 12

What is the net equation of the complex III reaction?

Answer 12

Question 13

What is cytochrome c’s prupose? structure? Qualities? Location?

Answer 13

Electron shuttle, carrying electrons from complex III to IV

Contains a iron heme group, is water soluble, and carries 1 electron total

Sits on the Inter-membrane space side of the IMM

Question 14

What is complex IV? What are the functions of complex IV?

Answer 14

Performs the final reduction of oxygen using electrons from cytochrome c

Prevents partially reduced oxygen (i.e. Reactive Oxygen Species, or oxygen radicals essentialy) from being released)

Pumps 2 electrons

Question 15

What is the structure of complex IV? What does it contain?

Answer 15

Contains 2 cytochromes, cytochromes a and a₃

Contains 2 copper centres; Cu_A and Cu_B

Oxygen binds between heme a₃ and Cu_B

Question 16

What is the net equation for the complex IV reaction?

Answer 16

Question 17

What are examples of toxic substances that block the ETC?

Answer 17

N₃ (azide), CO, CN^- (cyanide)

Question 18

What is F₀

Answer 18

One of the 2 ATP synthase subunits

Imbedded in IMM

Contains H+ channels

Subunits use ROMAN naming system

Question 19

What is F₁

Answer 19

Second ATP synthase subunit

Extends into matrix

Synthesizes ATP when linked to H+ in F₀

Subunits use GREEK naming system

Question 20

What is the structure of F₁ subunit

Answer 20

Has 3αß subunits arranged in a ball around the gamma subunit

Gamma unit spins from F₀ subunit activity, causing αß subunits to change between Open, Loose, and Tight site conformations

Open allows ATP and ADP to leave, Loose allows ADP and Pi to enter but not leave, and Tight sights creates ATP but retains it.

Cyclical action generates ATP

Question 21

How much ATP is synthesized per proton? Per NADH? FADH₂? What is the rationalle behind it?

Answer 21

4H+/ATP is best estimate;

3H+ to spin ATP synthase

1 H+ to move Pi

Thus:

10/4=2.5 NADH/ATP

6/4 = 1.5 ATP/FADH₂

Question 22

What is the structure of subunit c of F₀?

Answer 22

• Composed of 2 α-helices
• Span IMM
• 10-12 cylindrical shape
• 1/2 down one of the helixes is a Aspartic acid residue that can be protonated/deprotonated
• Entire subunit can rotate

Question 23

What is the structure of subunit a of F₀?

Answer 23

• Stationary
• Has a “clamp” that cover 2 of the c subunits
• Has 2 channels; one channel is open to the IMS and goes halfway down subunit a, while the other is open to the matrix and goes halfway up subunit a

Question 24

Which way does F0 spin?

Answer 24

Clockwise; such that proton goes through IMS 1/2 channel, forms uncharged aspartic acid, rotates around, and enters matrix 1/2 channel where subunit c deprotonates back into asparatate

Question 25

What is acceptor control?

Answer 25

The way phosphorylation of ATP is controlled: * when ATP/ADP ratio is High, Oxygen consumption drops (excess energy, ex. when sleeping) * when ATP/ADP ratio is low, oxygen consumption increases (lots of ADP to phosphorylate) \*ATP synthase cannot spin w/o ADP attaches to L state

Question 26

How can the ETC and ATP be chemically uncoupled? Provide an example and how it works

Answer 26

DNP (2,4-dinitrophenol) or FCCP Work by providing a way for H+ to be carried across IMM, lowering PMF ETC speeds up to replace lost H+; excess heat is produced but ATP synthesis slows down/remains the same (in simple terms, it creates a leak of protons)

Question 27

How can the ETC and ATP production be physiologically uncoupled? How?

Answer 27

Some tissues express UCP1 (uncoupling protein 1, AKA thermogenin) Transports H+ to matrix, bypassing ATP synthatse (similar to DNP, but regulated)

Question 28

What tissues express UCP1? What is it use? What are research implications of UCP1?

Answer 28

Brown fat expresses UCP1; important for thermoregulation (as by removing H+, ETC goes faster and produces heat) In obesiety, brown fat become dysfunctional → become white fat Research implication: white fat can be turned into "beige fat" (brown-like fat) w/ drugs, cold exposure, or excercise. Possible therapy for weight loss.

Question 29

Describe how the glycerol 3P shuttle works.

Answer 29

Cytosolic glycerol 3P dehydrogenase reduces DHAP to glycerol 3P, oxidizing NADH to NAD (glycerol 3P acts as electron carrier) IMM-bound glycerol 3P dehydrogenase reverses reaction, transfering the released electrons to FAD FAD passed to ubiquinone to form QH₂ QH₂ passes e to complex III \*Note this process is similar to that of complex II, so less H+ and thus less ATP yield

Question 30

What parts of the body uses the G3P shuttle for cytosolic NADH?

Answer 30

Skeletal muscle and the Brain

Question 31

What part of the body uses the Malate-aspertate shuttle?

Answer 31

Liver, Kidney, Heart

Question 32

Describe how the Malate-aspertate shuttle works

Answer 32

1. Oxaloacetate gets reduced into malate by malate dehydrogenase; malate carries NADH's electrons 2. Malate imported into matrix vis malate-α-ketoglutarate co-transporter 3. Malate dehydrogenase inside matrix converts malate back to oxaloacetate, transfering electrons to NADH (which goes to complex 1 of the ETC) 4. Oxaloacetate is transminated by aspartate aminotransferase, transfering amino group of glutamate to form α-ketoglutarate and aspartate. 5. Aspartate goes through glutamate-aspertate co-transporter, α-ketoglutarate goes through malate-α-ketoglutarate co-transporter to return to IMM 6. another aspartate aminotranferases undoes the reaction; turning aspartate back to oxaloacetate and α-ketoglutarate back into glutamate.

Question 33

How does ATP and ADP get transported across the IMM?

Answer 33

Via the adenine nucleotide translocase; an antiporter. Trades one ATP out from matrix for on ADP into matrix (from cytoplasm) Operates based of balance of charge (think Donnan potential, reaching charge equilbrium. Matrix is more negative than IMM due to pH build up)

Question 34

How does phosphate get tranpsorted across the IMM?

Answer 34

Via pyruvate translocase; pumps one H+ and one Pi into matrix. This is driven by the PMF, but decreases H+ gradient

Question 35

How does pyruvate get into the IMM?

Answer 35

SImilar to PI, using a symporter; specifically, Pyruvate translocase