10 - TCA and ETC

Question 1

What happens straight after pyruvate is formed?

Answer 1

• Pyruvate dehydrogenase (5 enzyme complex) oxidises pyruvate and adds CoA to form Acetyl CoA
• Occurs in mitochondrial matrix
• Irreversible loss of CO_{2 (regulatory step)}

Question 2

If you had a PDH deficiency, e.g due to genetics, what would happen?

Answer 2

Lactic acidosis, build up of pyruvate so would be converted to lactate by LDH to produce energy.

Question 3

Why is Vitamin B important?

Answer 3

Produces co-factors, e.g FAD and lipoic acid, that held PDH and therefore keep healthy metabolism

Question 4

What signals are PDH regulated by?

Answer 4

Question 5

How does the TCA cycle work?

Answer 5

2 TCA per glucose

6 NADPH

2 FADH₂

2 GTP

(2 CO₂)

Question 6

Summarise the key aspects of TCA

Answer 6

• In mitochondria
• Central pathway for sugars, FA, KB, AA, Alcohol
• Oxidative (Acetyl to 2CO₂)
• Produce intermediates that lose CO₂ easily to break C-C bonds
• Does not function in absence of O₂
• Provides precursors for biosynthesis
• No net synthesis of intermediates

Question 7

Where does regulation occur in TCA?

Answer 7

Isocitrate Dehydrogenase:

+ ADP - NADH, ATP (allosteric)

A-Ketoglutarate Dehydrogenase

• NADH, ATP, Succinyl CoA

Question 8

Give some examples of how TCA products act as biosynthetic precursors.

Answer 8

C5/C4 - AA

C4 - Haem and Glucose

C6 - FA

Question 9

Explain metabolism in terms of breaking bonds.

Answer 9

Breaking C-C bonds forming CO₂

Breaking C-H bonds and storing in NAD and FAD

Question 10

Where does most of the energy in catabolism come from?

Answer 10

NADH and FADH₂ transferring electrons to O_2, releasing large amounts of energy for ATP synthesis

Question 11

When intermediates are removed from TCA for biosynthesis, what replaces them?

Answer 11

• Breakdown of other molecules, e.g AA
• Pyruvate Carboxylase

Question 12

How many molecules of ATP are produced from each glucose and how many from each FADH₂ and NADH?

Answer 12

• 32
• 2.5 NADH
• 1.5 FADH₂

Energy released oxidation of these molecules used for oxidatitive phosphorylation

Question 13

Explain the electron transport chain up to just before ATP synthase

Answer 13

• 3 PTC
• NADH oxidised. 2 electrons pass to PTC and energy from this used to transport 2H+ across membrane.
• FADH oxidised at 2nd PTC
• Electrons passed to each PTC and energy used to move H+.
• Generates a proton motive force (electrochemical gradient)
• Oxygen is terminal electron acceptor and combines with 2 electrons and 2 H+ to form water

Question 14

How much of the energy from electrons is used to move H+ and what happens to the rest?

Answer 14

30%, rest of energy is lost as heat.

Tighter coupling, less energy loss to heat

Question 15

Explain how ATP is synthesis via oxidative phosphorylation?

Answer 15

• Oxidation of NADH set up p.m.f
• Protons move down their gradient via ATP synthase
• Energy from this dissapation is used to phosphorylate ADP

Question 16

How is oxidative phosphorylation regulated?

Answer 16

• High ATP, low ADP so no substrate
• Inward flow of H+ down ATP synthase stops
• Increase in H+ conc in intermembranal space
• Stops further H+ pumping and electron transport

Question 17

How is oxidative phosphorylation affected?

Answer 17

• Inhibitors (inhibition of electron transport, LETHAL, CN CO2)
• Uncoupling (FA and dinitrophenol+crestol) (no inhibition of electron transport)

- ATP concentration

- Genetic defects in proteins (decrease electron transport)

Question 18

How do inhibitors stop oxidative phosphorylation?

Answer 18

e.g CN

Block electron transport by binding to PTC, preventing oxygen binding so electrons can’t be terminally accepted

No p.m.f

Question 19

How does uncoupling affect oxidative phosphorylation?

Answer 19

• Uncouplers increase inner membrane permeability to H+
• Protons can diffuse down gradient wihout ATP synthase, removing p.m.f
• No p.m.f, no ATP synthesis, more energy lost to heat so get a temperature
• Uses up fat for fuel as metabolism is uncontrolled when uncoupled as no ATP high energy signals

Question 20

How does brown adipose tissue release heat?

Answer 20

• Contain thermogenin (UCP1)
• Noradrenaline released when cold
• Lipase stimuated, to release FA from TAG’s
• FA oxidised, producing NADH for electron transport
• Noradrenaloine activates UPC1
• UPC1 transport H+ back into mitochondria, energy from p.m.f not used for ATP synthesis so dissapates it as heat

Question 21

Who has brown fat?

Answer 21

Babies, mainly around their vital organs

Question 22

Compare oxidative and substrate level phosphorylation.

Answer 22

Oxidative has more efficient energy conversion so less energy lost to heat.

Substrate uses phosphorylate compound, not P_i

Question 23

What is the summary of ATP synthesis from glucose?

Answer 23

Question 24

What are the key points of electron transport and ATP synthesis?

Answer 24

• NADH and FADH₂ reoxidised and energy released from electrons as transfer down PTC
• O₂ required
• Large amounts of ATP produced

Question 25

What is dinitophenol and dinitrocresol and how do you treat them?

Answer 25

• Uncouples, found in slimming pills and herbicides
• Immediate referral to avoid death, e.g physical cooling, intravenous antioxidants like glutathione

Question 26

Describe all the stages of the Krebs Cycle.

Answer 26

Question 27

What are the functions of the TCA cycle?

Answer 27

Catabolic

• Oxidise acetyl of acetyl coA to 2 CO_2, GTP, NADH and FADH₂
• NADH and FADH₂ oxidised in electron transport chain to generate ATP by phosphorylation

Anabolic

- Provide intermediates for synthesis of haem, glutamate, aspartate etc

• Converting glucogenic amino-acids to glucose

Question 28

What are the consequences of a complete loss of enzyme in TCA cycle due to genetics?

Answer 28

• Baby would not survive
• Would make cycle inoperative so CNS and heart could not obtain enough energy to sustain life

Question 29

What are the key features of the electron transport chain?

Answer 29

• Electrons are transferred from NADH (and FAD2H) through a series of TPC to molecular oxygen with the release of free energy.
• The free energy is used to move protons from the inside to the outside of the inner mitochondrial membrane. The membrane itself is impermeable to protons and as electron transport proceeds the proton concentration on the outside of the inner membrane increases

Question 30

Why is cyanide toxic to our cells?

Answer 30

Binds to cytochrome site, stopping NADH and FADH being oxidised. Prevents generation of p.m.f and therefore ATP synthesis. No ATP then cell functions are impaired so cell death

Question 31

Bill, an agricultural worker, was employed by a farmer in East Anglia to spray 400 acres of wheat with a pesticide containing an aromatic weak acid, dinitrocresol (DNC). He failed to wear any protective clothing. After several days work, Bill fell ill with a very high temperature and profuse sweating. He was admitted to hospital, but in spite of strenuous attempts to decrease his body temperature, he fell into a coma and died. Autopsy showed a striking absence of subcutaneous fat.

Explain the symptoms.

Answer 31

Aromatic weak acids like 2-DNC (and 2,4-DNP) readily penetrate the mitochondrial inner membrane and act as uncoupling agents, i.e. they collapse the proton motive force and hence cause uncontrolled respiration to occur. This consumes large amounts of metabolic fuels (particularly fatty acids which are derived from fats stored in adipose tissue - hence the absence of subcutaneous fat) and of oxygen (this would lead to hypoxia but is prevented by increased pulmonary activity). As much less ATP than normal is made by oxidative phosphorylation under these conditions, a considerable amount of energy is lost as heat and the body temperature rises. When attempts to combat this by increased sweating eventually fail, the patient falls into a coma and dies.