MBC - Cell Integrity

Question 1

What are the reactions that occur in oxidative phosphorylation?

Answer 1

NADH + H+ + ½ O2 → NAD+ + H20

FADH2 + ½ O2 → FAD + H20

Question 2

What are the 4 membrane proteins (complexes) involved in the ETC?

Answer 2

1. NADH-Q oxidoreducatase or NADH reductase
2. Succinate-Q reductase or succinate dehydrogenase
3. Q-cytochrome C oxidoreductase
4. Cytochrome C oxidase

Question 3

What are the 2 mobile carriers of the ETC?

Answer 3

1. Co-enzyme Q (ubiquinone)

2. Cytochrome C

Question 4

Why does FADH2 generate less overall ATP than NADH?

Answer 4

FADH2 goes straight to complex 2 (succinate dehydrogenase), whereas NADH starts at complex 1 (NADH dehydrogenase).

Question 5

How does the ETC generate passive energy?

Answer 5

The movement of electrons from one complex to another via mobile carriers is energetically favourable, generating downstream energy which is then used to pump protons from carriers into the inter membrane space via complexes.

Question 6

How does the ETC generate ATP?

Answer 6

Protons flow down their concentration gradient via ATP synthase from the IM space into the matrix, thus generating ATP from ADP + Pi.

Question 7

Describe the function of ATP synthase in relation to its structure

Answer 7

ATP synthase is a multimeric enzyme consisting of an F0 bound subunit and an F1 unbound rotational unit. The way in which the subunit rotates is determined whether protons are flowing into or out of the matrix. The subunit rotation generates conformational change with altering affinities to either ATP or ADP.

Question 8

What do protons flowing into the matrix signify?

Answer 8

This means that subunit rotation induces and affinity to ADP, allowing for the generation of ATP

Question 9

What do protons flowing out of the matrix signify?

Answer 9

This means that subunit rotation induces an affinity to ATP for ATP hydrolysis.

Question 10

Describe the structure of an oxygen electrode

Answer 10

Measures the oxygen concentration of a solution housed within a small chamber. The base of the chamber is made of an oxygen permeable teflon membrane. Beneath the membrane there is a platinum cathode and a silver anode, along with an electrolyte (usually KCl)

Question 11

How does the oxygen electrode work?

Answer 11

Oxygen from the sample diffuses across the teflon membrane and reaches the platinum electrode. Here, it is reduced to from water, from the addition of electrons and protons. To complete the circuit, chloride from the electrolyte is reduced to silver chloride at the silver anode. We then read off the current from the ammeter, which is proportional to the oxygen concentration of a solution.

Question 12

What type of electrode do we use for measuring the ETC?

Answer 12

Clark-type oxygen electrode

Question 13

How do we measure the ETC?

Answer 13

Obtain a suspension of mitochondria from a tissue and place it in the chamber. We can then, measure the effects of various substrates or inhibitors on oxygen consumption over time.

Question 14

How do we measure basal respiration?

Answer 14

We take a reading of oxygen consumption after a minute to derive the basal rate of respiration.

Question 15

What happens when we add ADP to the electrode?

Answer 15

We will see a massive increase in oxygen consumption . If we know how much ADP is added and how much oxygen is consumed, we can work out the phosphorylation: consumption ratio = ADP-oxygen index for mitochondria efficiency.

Question 16

How do cyanide (CN-) and azide (N3-) act as metabolic posions?

Answer 16

They bind with high affinity to the ferric form of the haem group on complex IV (cytochrome oxidase)

Question 17

How does malonate act as a metabolic poison?

Answer 17

It has a similar structure to succinate therefore competitively inhibits succinate from transferring its electrons to succinate dehydrogenase (complex II) and then ubiquinone).

Question 18

How does rotenone act as a metabolic poison?

Answer 18

It is found in the roots and seeds of plants, and inhibits the transfer of electrons from complex I to ubiquinone.

Question 19

How does oligomycin act as a metabolic poison?

Answer 19

Antibiotic produced by streptomyces. It binds to the stalk of ATP synthase, thus blocking the movement of protons and thus limiting ATP synthesis from oxidative phosphorylation.

Question 20

How does DNP (dinitrophenol) act as a metabolic poison?

Answer 20

It is a proton ionophore which can shuttle protons across the inner mitochondrial membrane before reaching ATP synthase, thus uncoupling oxidative phosphorylation from ATP production.

Question 21

Why was DNP considered the ‘slimfast of the 1930s’?

Answer 21

It uncouples oxidative phosphorylation fro, ATP production, therefore making the body increase its metabolic rate (through oxidative phosphorylation) in order to produce the same amount of ATP as before. This increased metabolism (losing weight) but also increased body temperature.

Question 22

Why is DNP banned for diet pill use?

Answer 22

The margin between an effective dose of DNP for weight loss and a lethal dose was so slim that many people died as a result of DNP dosage. Although banned from diet pills, it is still found in pesticides and some food dyes.