9 - Metabolism 3 - oxidative phosphorylation

Question 1

how many electrons are produced in the reactions of glycolysis, pyruvate dehydrogenase complex, and the TCA cycle?

Answer 1

10 NADH
10 H+
2 FADH2

= each with 2 high energy electrons so total = 44

Question 2

what are electrons from NADH and FADH2 used for?

Answer 2

to reduce O2 and H2O = their energy is used to pump protons (H+) from mitochondrial matrix into intermembrane space

Question 3

what is pH environment comparison of intermembrane space & matrix?

Answer 3

pH decreased (acidic) in intermembrane space and increase in matrix

so protons flow back across the membrane following concentration gradient

energy of proton flow is used to phosphorylate ADP to ATP

Question 4

how doe NADH from cytoplasm get in?

Answer 4

with malate-aspartate shuttle

as NADH can’t directly cross inner mitochondrial membrane & can’t be re-oxidised to NAD+ directly using electron transport chain

Question 5

why is malate-aspartate shuttle important?

Answer 5

allows NADH into matrix - if no shuttle then metabolism would be uncoupled (glycolysis and oxidative phosphorylation separate)

Question 6

how does malate-aspartate shuttle work? (the malate part)

Answer 6

*so the last step in TCA cycle malate is converted to oxaloacetate and at the same time NAD+ makes NADH + H+

• this reaction can be reversed as well
• so in the intermembrane space the NADH that was made in glycolysis can be used to generate oxaloacetate to malate
• the malate-aspartate shuttle can then transfer malate to mitochondrial matrix
• the malate can then make conversion to oxaloacetate in TCA cycle which generate NADH in addition to malate that arises from fumarate

Question 7

what is the aspartate part of the malate-aspartate shuttle?

Answer 7

when the malate has been converted to oxaloacetate in matrix - some oxaloacetate used for TCA cycle, some of it is converted to aspartate (at the same time glutamate converted to alpha-ketoglutarate) and then the aspartate is taken through aspartate shuttle to cytosol where it can undergo reverse of reaction again to continue cycle of malate-aspartate shuttle

Question 8

what is the point of cycling of malate-aspartate shuttle?

Answer 8

to maintain redox balance between cytosol and mitchondria

Question 9

what is the electron transfer potential of NADH+ and FADH2 converted into?

Answer 9

the phosphoryl transfer potential of ATP

Question 10

what is phosphoryl transfer potential of ATP measured in?

Answer 10

measured by the free energy change, delta Go’, for the hydrolysis of ATP

Question 11

what is Electron transfer potential measured by?

Answer 11

the redox potential (or reduction potential), E’o, of a compound

Question 12

what is the standard redox potential E’o of a reduced substance X?

Answer 12

a measure for how readily X donates an electron (in comparison with H2)

Question 13

what does a negative and positive redox potential mean?

Answer 13

negative E’o = reduced form of X has a lower affinity for electrons than H2 (less likely to gain electrons)

positive means the opposite = higher affinity for electrons than H2 (more likely to gain electrons)

Question 14

what is the standard free energy change proportional to?

Answer 14

the change in standard redox potential and the number of electrons transferred

Question 15

what is the strongest of all terminal electron acceptors?

Answer 15

oxygen

Question 16

what makes up the respiratory chain?

Answer 16

group of proteins that site together
= complex 1,2,3,4

Question 17

how does respiratory chain start?

Answer 17

NADH donates a pair of electrons and the electrons allow complex 1 to pump protons (H+) from intracellular matrix intermembrane space

Question 18

what is complex 2 of respiratory chain? i.e. what does it couple as?

Answer 18

succinate dehydrogenase (which catalyses step 6 succinate - fumurate in TCA cycle)
= so electrons from FADH2

and also part of TCA cycle

Question 19

what are the proteins in respiratory chain linked by?

Answer 19

cytochrome C = takes electrons donated to complex 3 and links them to complex 4

Question 20

what are between complexes in respiratory chain?

Answer 20

electron carriers - ubiquinone, Q reductase & cytochrome C

Question 21

where do electrons from
a) NADH enter?
b) FADH2 enter?

Answer 21

a) at complex 1
b) at complex 2

Question 22

where are electrons ultimately transferred to in respiratory chain?

Answer 22

onto O2 to form H2O

Question 23

why are electrons handed to carriers in respiratory chain - why do they keep moving?

Answer 23

because they move to carriers with increasingly positive (oxidising) redox potentials

Question 24

describe the electrochemical gradient of mitochondria?

Answer 24

• more protons in intermembrane space than in matrix (due to being pumped out by complex 1,3,4)
• Forms an electrical field with the matrix side more negative
• Protons ‘want’ to flow back into the matrix
• Flow back into the matrix is coupled to ATP synthase which phosphorylates ADP +Pi -> ATP

Question 25

what is process of electron transport chain?

Answer 25

NADH passes electrons to complex 1 and FADH2 passes electrons to complex 2
- electrons from both complex 1 & 2 go to ubiquinone which pass electrons to cytochromes and to complex 3 then cytochrome C then complex 4 which transfers to O2 and makes H2O

Question 26

what can inhibit oxidative phosphorylation?

Answer 26

• carbon monoxide outcompetes oxygen (better at binding to haem groups so stops process of oxygen carriage and oxidative phosphorylation)
• cyanide & azide does the same

= they inhibit the transfer of electrons to O2 so no proton gradient formed and no ATP synthesised

Question 27

what is an example of uncoupling proteins?

Answer 27

thermogin in brown adipose tissue = brown adipose tissue contains a high number of mitochondria and is rich in uncoupling protein 1 (UCP1). UCP1 allows the uncoupling of oxidative phosphorylation from ATP synthesis, leading to the generation of heat without the production of ATP

non shivering thermogenesis - maintains body temp in newborns, hibernating animals cold-adapted mammals (so maintains temp when unable to shiver)

Question 28

what is uncoupling and what is the effect?

Answer 28

uncoupling = breaks the link between electron transport chain and ATP synthase (by inserting own protein channels - ionophores)

this means electrons bypass ATP synthase so protons go back into matrix instead so no proton gradient and no ADP→ ATP.

electrons still flow through complexes until end so oxygen keeps being delivered to tissue = as ADP rise, body increases metabolic rate to try increase NADH and FADH2 but useless →this all means more energy available to turn into heat energy

Question 29

what is P/O ratio?

Answer 29

• A measurement of the coupling of ATP synthesis to electron transport
• Number of molecules of inorganic phosphate (Pi) incorporate into ATP per atom of oxygen used
• Depends on the substrate which is oxidised

Question 30

what is P/O ratio of
a) NADH
b) FADH2

Answer 30

a) 1 NADH -> 3 ATP (as 3 proton pumps - 1,3,4)

b) 1 FADH2 -> 2 ATP (as only 2 proton pumps since skips complex 1 - proton pump 3,4 only)

Question 31

how many ATP does 1 glucose yield?

Answer 31

30-32 ATP (depends on precise values of P/O)