ATP and mitochondria

Question 1

where do oxidative phosphorylation reactions take place?

Answer 1

inner membrane

Question 2

where does the krebs cycle take place?

Answer 2

matrix

Question 3

why are the cristae folded?

Answer 3

increase SA for oxidative phosphorylation

Question 4

what is the equation for the reoxidation of NADH by molecular oxygen in mitochondria?

Answer 4

2NADH + 2H⁺ + O₂ —–> 2NAD⁺ + 2H₂O

Question 5

what is the equation for the reoxidation of FADH₂ by molecular oxygen in mitochondria?

Answer 5

2FADH₂ + O₂ —–> 2FADH₂ + 2H₂O

Question 6

what is ΔG for the reoxidation of NADH?

Answer 6

-223 kJ/mol

Question 7

what is ΔG for the reoxidation of FADH₂?

Answer 7

-170 kJ/mol

Question 8

what is the significance of ΔG for reoxidation of NADH and FADH₂?

Answer 8

ΔG for ATP hydrolysis is only -31 kJ/mol

energy released from reoxidation of NADH and FADH₂ enough to generate several phosphoanhydride bonds

much of this energy is recovered by components of ETC and used to synthesise ATP

Question 9

describe mtDNA’s genome.

Answer 9

16 569 bp circular genome

encodes only 37 genes

several copies can be found in one cell

Question 10

why is mtDNA important?

Answer 10

mutations in mtDNA is often a cause of genetic diseases

Question 11

how is mtDNA inherited and why is this important?

Answer 11

inherited via the ovum

therefore mtDNA mutations are transmitted to all maternal offspring

Question 12

describe the origins of the mitochondria.

Answer 12

occurred early in history of life on earth

evolutionary descendent of a prokaryote that established an endosymbiotic relationship with the ancestors of eukaryotic cells

many genes needed for mitochondrial function then translocated to the nuclear genome

Question 13

what are the 5 pieces of evidence that support the endosymbiont theory?

Answer 13

1 - mitochondria only arise from pre existing mitochondria

2- mitochondria have their own genome which resembles prokaryote genomes (circular molecule of DNA, no associated histones)

3- mitochondria have their own protein synthesising machinery, which resembles prokaryotic machinery rather than eukaryotic

4- the first amino acid of mitochondrial transcripts is fMet (formylated methionine residue), like in bacteria, rather that Met (methionine) like in eukaryotic proteins

5- many antibiotics (e.g streptomycin) that block bacterial protein synthesis also block mitochondrial protein synthesis but do not interfere with protein synthesis in eukaryotic cytoplasm

Question 14

what are the 3 enzymes involved in the ETC?

Answer 14

NADH dehydrogenase complex

cytochrome B-C1 complex

cytochrome oxidase complex

Question 15

what are the 2 carriers involved in the ETC?

Answer 15

ubiquinone (co enzyme Q)

cytochrome C

Question 16

what do the enzymes and carriers in the ETC do?

Answer 16

accept electrons and in doing so, a proton from the aqueous solution

as electrons pass through each of the complexes, a proton is pumped to the intermembrane space

Question 17

what is succinate dehydrogenase and what does it do?

Answer 17

integral membrane protein attached to inner surface of inner mitochondrial membrane

communicates directly with ubiquinone

As such one less proton is pumped to the intermembrane space, c.f. NADH and as a consequence, less ATP is produced.

The same is true for electrons donated by other FADH2 species, e.g. those generated in the glycerol phosphate shuttle and the β-oxidation of fatty acids.

Question 18

what are redox reactions?

Answer 18

electron transfer reactions

a reduced substrate donates electrons and becomes oxidised

an oxidised substrate/oxidant accepts electrons and becomes reduced

Question 19

what is a redox couple?

Answer 19

a substrate that exists in both oxidised and reduced forms

e.g NAD⁺/NADH
Fe³⁺/Fe²⁺
½O₂/H₂O

Question 20

what is the redox potential and how is standard E’0 determined?

Answer 20

ability to accept or donate electrons

determined experimentally

Question 21

what does a negative E’0 imply?

Answer 21

redox couple has a tendency to donate electrons

more reducing power than hydrogen

Question 22

what does a positive E’0 imply?

Answer 22

redox couple has a tendency to accept electrons

more oxidising power than hydrogen

Question 23

what does the transfer of electrons from one complex to another mean, energetically speaking?

Answer 23

transfer is energetically favourable

as electrons progress along the chain electrons lose energy

Question 24

what is ATP synthase?

Answer 24

multimeric enzyme

depending on proton flow direction, the complex either synthesises (protons move from intermembrane space to matrix) or hydrolyses (protons move from matrix to intermembrane space) ATP

Question 25

describe the structure of ATP synthase.

Answer 25

membrane bound part (F₀) part projects into the matrix (F₁)

Question 26

how does ATP synthase synthesise ATP?

Answer 26

rotation of the enzyme drives transition states, with altering affinities for ATP and ADP consequently, conformational energy flows from the catalytic subunit into the bound ADP and Pᵢ to promote the formation of ATP (chemical energy)

Question 27

describe the setup of an oxygen electrode.

Answer 27

small chamber within which the solution is housed base formed by oxygen-permeable teflon membrane under the membrane, a compartment contains the Pt cathode and the Ag anode

Question 28

what does an oxygen electrode do?

Answer 28

measures oxygen concentration in a solution

Question 29

how does an oxygen electrode work?

Answer 29

~0.6V applied between anode (+) and cathode (-) oxygen diffuses through the teflon membrane and is reduced to water at the Pt cathode O₂ + 4H⁺ + 4e⁻ ----> 2H₂O the circuit is completed at the Ag anode (oxidised to AgCl by the KCl electrolyte) 4Ag⁺ + 4Cl⁻ ----> AgCl + 4e⁻ the resulting current is proportional to the oxygen concentration in the sample

Question 30

how can the oxygen electrode by used to investigate the ETC?

Answer 30

suspension of mitochondria from a tissue placed into the oxygen electrode oxygen consumption of suspension monitored for set time period 1- baseline respiration measured 2- ADP added, causing rapid increase in oxygen consumption 3- if the quantity of ADP added is known then the ratio of the amount of ADP phosphorylated to the oxygen consumed can be calculated (ADP:oxygen index - measures efficiency of mitochondrial phosphorylation system) 4- ADP all consumed, system returns to basal respiration rate therefore, effects of various substrates and inhibitors on ETC can be determined

Question 31

why is any interruption to oxidative phosphorylation/ATP synthesis likely to cause cell death?

Answer 31

only a limited amount of ATP is available at any time each ATP molecule has a short lifespan (1-5 mins) therefore any interruptions causes rapid depletion and probably death, depending on cell type and metabolic requirements (only a few minutes for neurones, a few hours for muscle)

Question 32

what is the most common cause of a failure of oxidative phosphorylation?

Answer 32

lack of oxygen hypoxia/anoxia

Question 33

what is uptake of oxygen by mitochondria controlled by and why is it important?

Answer 33

controlled by components of ATP production (ADP and Pᵢ) respiratory control - allows body to adapt oxygen consumption to actual energy requirements

Question 34

what are metabolic poisons?

Answer 34

molecules that interfere with either the flow of electrons along the ETC or the flow of protons through ATP synthase, interrupting ATP synthesis highly toxic

Question 35

how do cyanide and azide act as metabolic poisons?

Answer 35

bind with high affinity to the ferric form (³⁺) of the haem group in the cytochrome oxidase complex blocks final step of ETC

Question 36

how does malonate act as a metabolic poison?

Answer 36

resembles succinate competitive inhibition of succinate dehydrogenase slows electron flow from succinate to ubiquinone by inhibiting oxidation of succinate to fumarate

Question 37

how does rotenone (isoflavone found in some roots and seeds) act as a metabolic poison?

Answer 37

inhibits transfer of electrons from complex I to ubiquinone

Question 38

how does oligomycin (antibiotic produced by Streptomyces) act as a metabolic poison?

Answer 38

inhibits oxidative phosphorylation binds to 'stalk' of ATP synthase blocks flow of protons through the enzyme

Question 39

how does dinitrophenol (DNP) act as a metabolic poison

Answer 39

proton ionophone shuttles protons across inner mitochondrial membrane

Question 40

how can DNP induce weight loss?

Answer 40

transports protons across the mitochondrial membrane this uncouples oxidative phosphorylation from ATP production leads to an increase in metabolic rate and body temperature

Question 41

why was DNP abandoned as a weight loss drug?

Answer 41

margin between effective and lethal doses is very slight many deaths and permanent injuries as a result now illegal to sell for human consumption, although people still buy and use it with painful consequences

Question 42

where is DNP still used?

Answer 42

pesticides food dye