ATP and mitochondria Flashcards
where do oxidative phosphorylation reactions take place?
inner membrane
where does the krebs cycle take place?
matrix
why are the cristae folded?
increase SA for oxidative phosphorylation
what is the equation for the reoxidation of NADH by molecular oxygen in mitochondria?
2NADH + 2H⁺ + O₂ —–> 2NAD⁺ + 2H₂O
what is the equation for the reoxidation of FADH₂ by molecular oxygen in mitochondria?
2FADH₂ + O₂ —–> 2FADH₂ + 2H₂O
what is ΔG for the reoxidation of NADH?
-223 kJ/mol
what is ΔG for the reoxidation of FADH₂?
-170 kJ/mol
what is the significance of ΔG for reoxidation of NADH and FADH₂?
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
describe mtDNA’s genome.
16 569 bp circular genome
encodes only 37 genes
several copies can be found in one cell
why is mtDNA important?
mutations in mtDNA is often a cause of genetic diseases
how is mtDNA inherited and why is this important?
inherited via the ovum
therefore mtDNA mutations are transmitted to all maternal offspring
describe the origins of the mitochondria.
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
what are the 5 pieces of evidence that support the endosymbiont theory?
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
what are the 3 enzymes involved in the ETC?
NADH dehydrogenase complex
cytochrome B-C1 complex
cytochrome oxidase complex
what are the 2 carriers involved in the ETC?
ubiquinone (co enzyme Q)
cytochrome C
what do the enzymes and carriers in the ETC do?
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
what is succinate dehydrogenase and what does it do?
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.
what are redox reactions?
electron transfer reactions
a reduced substrate donates electrons and becomes oxidised
an oxidised substrate/oxidant accepts electrons and becomes reduced
what is a redox couple?
a substrate that exists in both oxidised and reduced forms
e.g NAD⁺/NADH
Fe³⁺/Fe²⁺
½O₂/H₂O
what is the redox potential and how is standard E’0 determined?
ability to accept or donate electrons
determined experimentally
what does a negative E’0 imply?
redox couple has a tendency to donate electrons
more reducing power than hydrogen
what does a positive E’0 imply?
redox couple has a tendency to accept electrons
more oxidising power than hydrogen
what does the transfer of electrons from one complex to another mean, energetically speaking?
transfer is energetically favourable
as electrons progress along the chain electrons lose energy
what is ATP synthase?
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
describe the structure of ATP synthase.
membrane bound part (F₀)
part projects into the matrix (F₁)
how does ATP synthase synthesise ATP?
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)
describe the setup of an oxygen electrode.
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
what does an oxygen electrode do?
measures oxygen concentration in a solution
how does an oxygen electrode work?
~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
how can the oxygen electrode by used to investigate the ETC?
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
why is any interruption to oxidative phosphorylation/ATP synthesis likely to cause cell death?
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)
what is the most common cause of a failure of oxidative phosphorylation?
lack of oxygen
hypoxia/anoxia
what is uptake of oxygen by mitochondria controlled by and why is it important?
controlled by components of ATP production (ADP and Pᵢ)
respiratory control - allows body to adapt oxygen consumption to actual energy requirements
what are metabolic poisons?
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
how do cyanide and azide act as metabolic poisons?
bind with high affinity to the ferric form (³⁺) of the haem group in the cytochrome oxidase complex
blocks final step of ETC
how does malonate act as a metabolic poison?
resembles succinate
competitive inhibition of succinate dehydrogenase
slows electron flow from succinate to ubiquinone by inhibiting oxidation of succinate to fumarate
how does rotenone (isoflavone found in some roots and seeds) act as a metabolic poison?
inhibits transfer of electrons from complex I to ubiquinone
how does oligomycin (antibiotic produced by Streptomyces) act as a metabolic poison?
inhibits oxidative phosphorylation
binds to ‘stalk’ of ATP synthase
blocks flow of protons through the enzyme
how does dinitrophenol (DNP) act as a metabolic poison
proton ionophone
shuttles protons across inner mitochondrial membrane
how can DNP induce weight loss?
transports protons across the mitochondrial membrane
this uncouples oxidative phosphorylation from ATP production
leads to an increase in metabolic rate and body temperature
why was DNP abandoned as a weight loss drug?
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
where is DNP still used?
pesticides
food dye
