Barrow: Citric Acid Cycle and Terminal Respiration

Question 1

What is the main input to the citric acid cycle? Output?

Answer 1

Acetyl CoA, CO₂ + energy

Question 2

What is an amphibolic cycle?

Answer 2

Builds things up and breaks them down (like the TCA cycle)

Question 3

Other names for the Citric Acid Cycle?

Where does it occur?

What fuel molecules enter it?

Answer 3

Krebs cycle, TCA (tricarboxylic acid) cycle

Mitochondrial matrix (inside inner membrane)

All (carbs, fatty acids, and amino acids)

Question 4

TCA cycle: summarise its general purpose

In collaboration with oxidative phosphorylation, how much of aerobic cell energy is made through the TCA cycle?

Does it directly produce ATP?

Answer 4

Removes electrons from molecules and passes them onto electron carriers (NAD+, FADH+)

90%

No

Question 5

Walk through the evolution of the TCA cycle

Answer 5

Plants increased atmospheric oxygen (which then causes free radicals, oxidises organic molecules, etc) -> primitive organisms evolved to use oxygen to oxidise food molecules beyond glycolysis, giving them much more energy, and an evolutionary advantage

Question 6

Give an overview of everything, from glycolysis to terminal respiration

Answer 6

Question 7

Why does oxygen (alongside TCA cycle and terminal respiration) mean you no longer need the glycolytic pathway to regenerate NADH?

Answer 7

Because NADH (and other electron carriers) can now, through many steps, donate that extra H ion to oxygen instead (re-oxidising it to water)

Question 8

What converts into Acetyl CoA?

Where does this occur?

Answer 8

Pyruvate from glycolysis and fatty acid breakdown

Mitochondrial matrix (within inner membrane)

Question 9

Describe overview of process from when pyruvate enters the mitochondrial matrix [picture]

Answer 9

Question 10

What enzyme converts pyruvate to Acetyl CoA?

Answer 10

Pyruvate dehydrogenase

Question 11

Pyruvate dehydrogenase: how big? How many subunits (and how many copies of each?)

Answer 11

50nm across, 3 subunits (10 copies)

Question 12

How many carbons enter the TCA cycle as acetyl CoA? What form do they leave as?

Answer 12

2 carbons, CO₂

Question 13

Complex version of TCA cycle (not for revision, but just to see if you can follow it)

Answer 13

Question 14

What is pyruvate dehydrogenase (pyruvate to Acetyl CoA) regulated by?

Answer 14

Inhibited by Acetyl CoA, NADH and ATP. Boosted by pyruvate and ADP.

Question 15

What two points within the TCA cycle can it also be regulated?

Answer 15

Isocitrate dehydrogenase (+: ADP, -: ATP & NADH) and alpha-ketoglutarate dehydrogenase (-: ATP, succinyl CoA, NADH)

Question 16

TCA cycle control checkpoints (isocitrate dehydrogenase and alpha-ketoglutarate dehydrogenase): how does this allow for re-direction of cellular resources?

Answer 16

Isocit: causes citrate build up (as they are interconvertible), which shuttles back to cytoplasm, and causes phosphofructokinase to stop glycolysis

Alpha: causes build up of alpha-ketoglutarate when enzyme is inactive -> production of amino acids

Question 17

Just another image to appreciate - how things are converted through the amphibolic TCA cycle

Answer 17

Question 18

What problem stems from the fact that the TCA cycle intermediates can be used for other purposes? Give an example of how his can occur.

What process reverses the example you gave? What is this reaction known as?

Answer 18

If they are used too much, then the TCA cycle can shut down (such as when exercising muscle makes ATP and uses up oxaloacetate)

Gluconeogenesis pathway: pyruvate being converted to oxaloacetate by pyruvate carboxylase (which is activated by acetyl CoA, a build up of which would demonstrate the TCA cycle is not happening) - anaplerotic reaction (“to fill up”)

Question 19

What is the name of the cycle that plant (especially seeds) use to convert stored fats into carbohydrates? What is the cellular organelle called, and where does it sit?

Answer 19

Glyoxylate cycle (part of TCA with two new enzymes involved) - occurs in glyoxysome (next to lipids + mitochondria)

Question 20

Describe a mitochondria

Answer 20

Matrix -> inner membrane -> intermembrane space -> outer membrane

Question 21

Where are the majority of reduced electron carriers formed?

NADH cannot cross membranes of mitochondria. How do electrons gained from glycolysis get into the mitochondria? Describe.

Answer 21

Mitochondrial matrix

Glycerol phosphate shuttle (NADH passes electrons to DHAP [see glycolysis] converting it to Glycerol-3-Phosphate -> binds with enzyme on outer membrane, which can pass electrons to FAD, which then enters electron transport chain)

Question 22

How many complexes make up the electron transport chain (bonus points for names)? Where do NADH electrons enter? What about FADH₂?

Answer 22

NADH-Q Oxioreductase, Succinate-Q Reductase, Q-cytochrome c oxioreductase, cytochrome c oxidase

Question 23

Terminal respiration complex 1: what is it called? [clue: what feeds in? What does it pass stuff to?]

What is used to carry electrons through? What are they passed to, and what does this create?

Answer 23

NADH-Q Oxidoreductase: utilises Iron-Sulphur centres and flavin mononucleotide to pass electrons to ubiquinone (Q), which is converted to ubiquinol (QH₂)

Question 24

What do amytal (barbituate), piercidin A (antibiotic), and rotenone (insectiside) have in common?

Answer 24

Inhibit electron flow from Fe-S centres of terminal respiration complexes to ubiquinone (blocks electron transport chain)

Question 25

Terminal Respiration Complex 2: what is it called? [Clue: also part of citric acid cycle - is a smaller part of this larger enzyme] What feeds in? What transfers electrons, and to what? How does it differ from complex 1?

What does it have in common with blood? What is it used for?

Answer 25

Succinate-Q Reductase: FADH₂ passes electrons to Fe-S centres to ubiquinone (-> ubiquinol, QH₂) - unlike complex 1, it doesn’t transport H+ into intermembrane space.

Has a heme group - blocks stray electrons and stops them from becoming free radicals

Question 26

What is Q₁₀ also known as?

Answer 26

Ubiquinone (NB: in skin cream, but it’s all bullshit as the concentration isn’t high enough)

Question 27

What gets passed on from complex 1 to complex 3? What is complex 3 called? What is its purpose?

Answer 27

Q-cytochrome c oxioreductase passes electrons from one Ubiquinone (QH₂) molecule -> two cytochrome c molecules (and pumps protons into intermembrane space)

Question 28

What shuttles between terminal respiration complex 3 and 4? What is the complex called? What are electrons passed through/to? Does it also pump protons?

Why are there heme groups?

Answer 28

Cytochrome c (to cytochrome c oxidase): electrons channelled through Fe-Cu centres to oxygen. Protons are also pumped

Purpose of heme groups: channel electrons, reduce free radicals (same as in other complexes)

Question 29

Another nice picture: the entire terminal respiration chain. NADH and FADH2 are oxidised, with their electrons passed to QH2. QH2 passes (it is oxidised) the electrons to cytochrome c (it is reduced) in complex 3, and cytochrome c then passes (becoming oxidised) them on to complex 4. Protons are pumped in complexes 1, 3, and 4

Answer 29

Question 30

What is chemiosmosis (in reference to terminal respiration)?

Why is this vectoral?

What do the protons act as? What is it called when they are released to do work?

Answer 30

Protons passing from matrix to intermembrane space as electrons move through complexes of transport chain.

Because the reactions have particular spatial directionality

Store of potential energy. Proton motive force is when they flow back down their gradient and do work

Question 31

What is the name of the protein that protons flow from the space back into matrix through? Describe roughly what it does…

Answer 31

ATP synthase (ATPase) - has a central pore that allows protons to pass through, and the energy is used to convert ADP+P to ATP and kick the ATP out…

Question 32

ATP synthase: how many parts? What do they do?

Answer 32

2 parts (F₀ = membrane bound proton conducting unit [10 subunits] w/separate subunit to connect to… F₁ = protrudes into matrix and acts as catalyst for ATP synthesis)

Question 33

ATP synthase: lay out the steps of its operation

Answer 33

ADP + P enters beta subunit of F1. Rotation of F0 and gamma shaft forces conformational change of beta subunit, catalyzing conversion of ADP+P -> ATP.

Question 34

ATP synthase: what is the largest amount of energy used for?

Answer 34

Pushing ATP from beta subunit into matrix

Question 35

What is the mechanism of ATP synthase called? What is involved?

Answer 35

Binding change mechanism: protons moving from positive to negative side of membrane causes sequential changes of beta subunits (one that binds ADP/P, one binding ATP, one that binds neither)

Question 36

What experimental evidence is there for this mechanism of ATP synthase in action?

Answer 36

Can take F1 portion with an actin filament (w/fluorescent compounds) attached to the gamma subunit, attach the whole thing to a coverslip, and feed it ATP - then you can watch it spin round as it reverses its usual process

Question 37

Why is it called oxidative phosphorylation?

Answer 37

You are oxidising NADH and FADH2, and then using that energy to phosphorylate ADP -> ATP

Question 38

How many mol of ATP are generated per mol of NADH and FADH2 respectively? Why?

Answer 38

2.5 and 1.5 - electrons moving through all four terminal respiration complexes move 8 protons across, and ATP synthase produces one ATP for every 3 protons it shuttles back into the matrix. FADH2 only enters at complex 2, though, so less protons are pumped across (and less ATP is generated).

Question 39

How many protons do all four terminal respiration complexes move across the inner mitochondrial membrane? How many protons does it cost to create 1 ATP?

Answer 39

8, 3

Question 40

How many ATP are made through glycolysis, through pyruvate oxidation, and through Acetyl-CoA oxidation? And in total?

Answer 40

Question 41

Electron transport is said to be _______ to ATP synthesis. If _________, you can get _________ ___________. What is going on?

Answer 41

coupled, uncoupled, malignant hypothermia

Inner mitochondrial membrane becomes permeable to proteins, which means gradient cannot be generated - electron transport still occurs, with oxygen being reduced to water, but no ATP is made (process is “uncoupled). The energy is instead released as heat.

Question 42

What happens if you get malignant hypothermia?

Who does this happen to?

Answer 42

Large increase in body temperature. Muscle cells become irreversibly damaged from excessive heat build up

Susceptible people exposed to halothane (or halothane-like drugs) [1 in 75000 people, 1 in 15000 kids]

Question 43

When do you get intentional uncoupling of the electron transport chain from ATP synthase?

How does this process work?

Answer 43

Brown fat in newborn infants (and other mammals) - it’s brown because it’s jam packed with mitochondria, which have deoxygenated heme groups as part of most of the electron transport complexes

When creature becomes cold, norepinephrine triggers opening of channel in protein called thermogenin (which sits on inner mitochondrial membrane of brown fat cells, causes uncoupling of ETC from ATP synthase, and generates heat

[NB: plants do it too - arum lily to attract insets, skunk cabbage to melt snow, etc]