Title: BV2 / BV2 - 2 / Modul 5 / Inside the mitochondria

Q: How is the pyruvate dehydrogenase complex regulated?
A: - Phosphorylation of E1 by Pyruvate dehydrogenase kinase inhibits PDH
- PDH phosphatase activates E1
- Ca2+ increases PDH activity
- Acetyl-CoA is an allosteric inhibitor of E2
- NADH is an allosteric inhibitor of E3

Q: What affects pyruvate dehydrogenase activity?
A: Ca2+ increases PDH activity

Q: How can E1 be regulated?
A: - Activated by PDH phosphatase
- Inhibited by phosphorylation of E1 via PDH kinase

Q: How can E2 be regulated?
A: Can be inhibited by the allosteric inhibitor acetyl-CoA

Q: How can E3 be regulated?
A: Is inhibited allosterically by NADH

Q: What happens in the 8 steps of the Krebs cycle?
A: 1. Condensation by citrate synthase - needs H2O
2. Restructuring by aconitase
3. Oxidation by isocitrate dehydrogenase - release of CO2 and NADH
4. Oxidation by alfa-ketoglutarate dehydrogenase - release of CO2 and NADH
5. Release of CoA is coupled to GTP synthesis by succinyl-CoA synthase
6. Succinate dehydrogenase oxidizes succinate via FAD to reduce O
7. Fumarase hydrates
8. Malate dehydrogenase forms oxaloacetate forming NADH

Q: What happens in step 1 of the TCA cycle?
A: 1. Condensation by citrate synthase - needs H2O

Q: What happens in step 2 of the TCA cycle?
A: 2. Restructuring by aconitase

Q: What happens in step 3 of the TCA cycle?
A: 3. Oxidation by isocitrate dehydrogenase - release of CO2 and NADH

Q: What happens in step 4 of the TCA cycle?
A: 4. Oxidation by alfa-ketoglutarate dehydrogenase - release of CO2 and NADH

Q: What happens in step 5 of the TCA cycle?
A: 5. Release of CoA is coupled to GTP synthesis by succinyl-CoA synthase

Q: What happens in step 6 of the TCA cycle?
A: 6. Succinate dehydrogenase oxidizes succinate via FAD to reduce O

Q: What happens in step 7 of the TCA cycle?
A: 7. Fumarase hydrates

Q: What happens in step 8 of the TCA cycle?
A: 8. Malate dehydrogenase forms oxaloacetate forming NADH

Q: In which 3 ways can regulation of Krebs occur?
A: 1. Citare synthase by feedback inhibition
2. Isocitrate is irreversible and therefore regulated
3. Alfa-ketoglutarate dehydrogenase is irreversible and is a major pont of replenishing the TCA cycle via glutamine/glutamate

Q: What can citrate synthase be activated and inhibited by respectively?
A: Activated
+ ADP
Inhibited
- NADH
- ATP
- Citrate
- Succinyl-CoA

Q: What can isocitrate dehydrogenase be activated and inhibited by respectively?
A: Activated
+ ADP
+ NAD+
+ Ca2+
Inhibited
- ATP
- NADH

Q: What can the alfa-ketoglutarate dehydrogenase complex be activated and inhibited by respectively?
A: Activated
+ Ca2+
Inhibited
- NADH
- ATP
- GTP
- Succinyl-CoA

Q: What in the TCA is activated by Ca2+?
A: - Isocitrate dehydrogenase
- Alfa-ketoglutarate dehydrogenase complex

Q: What is the TCA is inhibited by succinyl-CoA?
A: - Citrate synthase
- Alfa-ketoglutarate dehydrogenase complex

Q: How does the food we eat leave our body?
A: U breathe out the food u eat

Q: What is the molecular formula for the oxidation of glucose?
A: C6H12O6 + 6O2 -> 6CO2 + 6H2O + heat

Q: What is DG of oxidation of glucose?
A: DG = -2880 kJ/mol of glucose

Q: Which reactions can the oxidation of glucose be divided into?
A: 1. Glycolysis
2. The TCA cycle
3. Oxidative phosphorylation

Q: What is the net of products produced in glycolysis and Krebs?
A: Glucose -> 10 NADH + 2 FADH2 + 4ATP

Q: What is the chemiosmotic theory?
A: The reduction of oxygen is coupled to the transfer of protons across a semipermeable membrane
- The energy stored in this gradient is used to synthesize ATP

Q: What is the proton motif force equal to?
A: Membrane potential + pH gradient = proton motif force

Q: What is the redox potential?
A: The redox potential is the likelyhood of a chemical compound to accept or donate electrons

Q: What does a positive redox potential indicate?
A: Positive - how likely u are to be in the reduced state

Q: What does a negative redox potential indicate?
A: Negative - how likely u are to be in the oxidated state

Q: What is the redox potential (Em) for NAD+/NADH?
A: - 320mV

Q: What is the redox potential (Em) for O2/H2O?
A: +820mV

Q: Whic complexes does the respiratory chain consist of?
A: complexes 1-4

Q: What is the 5th complex in the ETC?
A: ATP synthase

Q: What does the first motor in ETC do?
A: Oxidises NADH

Q: Where does oxidation of NADH happen in ETC?
A: Happens in complex I

Q: What are the complexes, I, III, and VI involved in doing in the ETC?
A: Are all involved in pumpung H+ over the memrane into the matrix

Q: What happens during the transfer of the electrons between the complexes in the ETC?
A: During the transfer of the electrons between the complexes we get an electrial transmembrane potential

Q: What does complex 2 do in the ETC?
A: Oxidise FADH2 into FAD
- Succinate dehydrogenase - from TCA

Q: What is the functional process in bioenergetics?
A: The transfer of e- from one redox reaction to another

Q: How many electron carriers are there in the ETC?
A: More than 20

Q: What is the reaction of the complexes in the ETC that oxidise NADH?
A: NADH + 1/2 O2 -> NAD+ + H2O

Q: What is electron flow in the ETC influenced by?
A: - Distance of 2 redox reactions
- Redox potential
- Response to loss/gain of an e- of the donor/acceptor

Q: What potential of NADH will we start with in the respiratory chain (ETC)?
A: Start with a low redox potential of NADH

Q: What does NAD+ stand for?
A: Nicotinamide adenine dinucleotide +

Q: What is transfered in the reduction of NAD+ to NADH?
A: 2H+ + 2e-

Q: Which are the 3 mobile transport carriers in the ETC?
A: 3 mobile transport carriers in ETC
- NAD+
- Ubiquinone
- Cytokrome c

Q: What is another name of ubiquinone?
A: Co-enzyme Q

Q: What is ubiquinone?
A: A small, fat-soluble redox co-factor
- 1 electron unstable state - ubiquinone

Q: What is ubiquinones structure?
A: A benzoquinone ring with a lipid soluble polypernoid side chain

Q: How does ubiquinone change when taking up electrons?
A: Ubiquinone - (+ e-) -> semiquinone (1 e-) - (+ e-) -> ubiquinol (2e-)

Q: What are examples of free co-factors of electron transfer?
A: - Ubiquinone
- Cytochrome C

Q: What is cytochrome C?
A: Single electron carrier

Q: What can be said about cytochrome C's properties and structure?
A: It's water soluble and contains a single c-type heme
- Is a single electron carrier

Q: What are examples of bound co-factors of electron transfer?
A: - Heme (cytochromes: 1e-)
- Fe-S clusters (1e-)
- FADH2 (2e-)

Q: What is Heme (cytochromes: 1e-) classified by?
A: According to the structure of the porphyrin prosthetic group
- E.g. a-, b-, c-type heme

Q: What are Fe-S clusters?
A: Fe inons around a sulfate
- Can carry 1e-

Q: How can FADH2 be described briefly?
A: Covalently bound as FAD or FMN
- Carries 2 electrons

Q: What does FAD stand for?
A: Flavin adenine dinucleotide

Q: What type of ring structure does FADH2 have attached?
A: An isoalloxazine ring

Q: What does the 1st complex in the ETC do?
A: - Oxidises NADH to NAD+
- Reduces ubiquinone to ubiquinol

Q: What's the structure of the complex 1 (ETC)?
A: Large, multi-subunit complex of almost 1MDa

Q: How many of the complex 1 are mtDNA enconded (ETC)?
A: 7 are mtDNA encoded

Q: When can one have reduced activity in complex 1 of ETC?
A: - Reduced activity with age
- Reduced activity in Parkinson's disease

Q: What is complex 1 of the ETC involved in?
A: Involved in ROS formation (reactive oxygen species)

Q: What does ROS stand for?
A: Reactive oxygen species

Q: Which enzymes can be found in complex 1?
A: - NADH dehydrogenase
- NADH ubiquinone oxireductase

Q: Which enzymes can be found in complex 2?
A: - Succinate dehydrogenase
- Succinate ubiquinone oxidoreductase

Q: What is the molecular formula of the complex 1 reaction?
A: NADH + H+ + CoQ + 4H+(in) -> NAD+ + CoQH2 + 4H+(out)

Q: What is the molecular formula of the complex 2 reaction?
A: Succinate + CoQ -> fumarate + CoQH2

Q: What does complex 2 in ETC consist of?
A: - 4 subunits
- Contains an FAD moiety (fragment of molecule)

Q: What does complex 2 do in ETC?
A: - Oxidises succinate to fumarate
- Reduces ubiquinone to ubiquinol
- Links TCA cycle to oxidative phosphorylation

Q: What does not occur in complex 2 of the ETC?
A: No prroton translocation

Q: What is the molecular formula of the complex 3 reaction?
A: QH2 + 2Fe3+-Cyt c ----> Q + 2Fe2+-Cyt c + 2H+

Q: Which enzymes are active in complex 3 (ETC)?
A: - Ubiquinol:cytochrome c oxidoreductase or bc1-complex

Q: What does complex 3 contain?
A: Contains 1 FeS cluster (Rieske protein)

Q: What does complex 3 in ETC do?
A: - Oxidises ubiquinol to ubiquinone
- Reduces cytochrome c
Transfer from 2e- to 1e- carrier, which requires an intermediate step of semi-ubiquinone

Q: What are other non-membrane bound ubiquinone oxidoreductases?
A: - ETFDH: electron transfer flavin dehydrogenase
- GPDH: glycerol-3-phosphate dehydrogenase
- DHODH: dihydroorotate dehydrogenase

Q: When is ETFDH: electron transfer flavin dehydrogenase used?
A: In beta-oxidation

Q: When is GPDH: glycerol-3-phosphate dehydrogenase used?
A: In glycerol metabolism

Q: When is DHODH: dihydroorotate dehydrogenase used?
A: In pyrimidine biosynthesis

Q: What does complex 4 contain?
A: Contains 2 hemes (type a) and 2 copper centres

Q: Which enzymes are active in complex 4?
A: - Cytochrome c oxidase
- Cytochrome c oxidoreductase

Q: What is the molecular formula for the complex 4 reaction?
A: 4Fe2+-cytochrome c + 8H+(in) + O2 -> 4fe3+-cytochrome c + 2H2O + 4H+(out)

Q: What does complex 4 do in the ETC?
A: Collects 4 e- to transfer to O2, forming water

Q: What is complex 4 sensitive to?
A: Cyanide

Q: What are reactive oxygen species (ROS) important for?
A: - Cell signalling
- Homeostasis
- Immune response
- Wound healing

Q: What are reactive oxygen species (ROS) damaging in?
A: - Cancer
- Ischaemia
- Oxidative stress
- Apoptosis

Q: What is a free radical?
A: An unpaired electron in the outer orbital

Q: What is the reaction process/formula for ROS?
A: O2 (+ e-) superoxide, O2- (+ e-) hydrogen peroxide, H2O2 (+ e-) Hydroxyl radical, OH (+ e-) H2O

Q: What does complex 5 consist of?
A: 16 different subunits
- ATP6 and ATP8 are mtDNA encoded

Q: What is the structure of complex V?
A: - Water soluble F1 fraction contains active site (outside membrane)
- F0 fraction (inside cell) forms a ring structure within the membrane and contains the proton channel

Q: Which enzymes are active in complex V?
A: - ATP synthase
- F0F1 ATPase

Q: What does the F0 fraction consist of?
A: This complex has a ring of spinning subunits
- In the middle of the c-subunit is a gamma stalk, which makes it spin and in each c-subunit there is one Aspartate 61
- a and b-subunits can be opened or closed

Q: What causes c-subunits to rotate?
A: Rotation of c-subunits is caused by H+

Q: What happens when the H+ has gone around the ring?
A: When H+ was gon all aorund the ring it comes to a channel and it wants to go into the matrix

Q: How many rotations of the ring of spinning c-subunits are there per sec?
A: 200 rotations/sec

Q: How amny ATPs are made everytime 1 ring goes around?
A: 3 ATP everytime 1 ring goes around

Q: How does the F0 subunit work?
A: - Aspartate + H+ -> aspartic acid (more hydrophobic)
- The c subunit ring rotates until the aspartic acid is again exposed to the lower half-channel of the a-subunit
- The H- enters the matrix

Q: How does the F1 fraction work?
A: - The gamma stalk rotates with the c-ring, while the b-subunit fixates the a-, alfa-, and beta-subunits
There are 3 phases:
- Open (free)
- Loose (ADP + Pi bound)
- Tight (ATP-bound)
- 1 rotation of the c ring results in 3 ATP

Q: How many times is each ATP in our body used?
A: Each ATP in our body is used around 500 times

Q: How much ATP do we use per day?
A: We use the equivalent of our own weight in ATP every day

Q: What does the inner mitochondrial membrane consist of?
A: - High in phospholipids
- Cardiolipids, phosphatidylcholine, etc.

Q: What is the protein/lipid ratio in the inner mitochondrial membrane?
A: 80/20

Q: What is the inner mitochondrial membrane impermeable to?
A: Impermeable to all but O2, CO2, and H2O

Q: What can generally be said abou the transporters of the inner mitochondrial membrane?
A: - Often highly specific
- Can be regulated

Q: Which typer of transporters can be found in the inner mitochondrial membrane?
A: - Protonophore
- Ionophore
- Uniporter
- Symporter
- Antiporter

Q: What do uncoupling proteins do?
A: Uncoupling proteins create a channel, alowing protons (H+) to reenter the mitochondrial matrix without capturing any energy as ATP

Q: What does uncoupling of respiration from ATP synthesis do?
A: - Regulation of ATP synthesis rate
- Heat production in BAT (UCP1)
- Regulation of ROS (UCP2)
- Function of UCP3 is unknown

Q: How many uncoupling proteins are there?
A: - Heat production in BAT (UCP1)
- Regulation of ROS (UCP2)
- Function of UCP3 is unknown

Q: What happens to the used protons after ATP synthesis?
A: The protons do not reenter the matrix again by any means

Q: What does brown adipose tissue do?
A: Brown adipose tissue is there to generate heat
 allows H+ to reenter the matrix not via a complex
- Not inhibited by complex 5

Q: What does UCP1 allow for?
A: Allows H+ to reenter the matrix, without capturing any energy as ATP
- However, this generates heat

Q: How can the ETC be summarised?
A: The transfer of electrons to oxygen is coupled to the transfer of protons across a semipermeable membrane. The electrochemical force (membrane potential) can be used for e.g. substrate import/export, ATP synthesis and heat production

Q: What uses the most O2 in the cell?
A: Mitochondria

Q: Where is most O2 used in the mitochondria?
A: O2 consumption is coupled to ATP synthesis

Q: What happens to the oxgygen we breathe in, how does it exit our body?
A: We pee out the oxygen

Q: WHat is all the oxygen we use used for?
A: All the oxygen we use is used for ATP

Q: What is the name of complex 1 in ETC?
A: NADH dehydrogenase

Q: What is the name of complex 2 in ETC?
A: Succinate dehydrogenase

Q: What is the name of complex 3 in ETC?
A: CoQ-Cytochrome-b-oxioreductase

Q: What is the name of complex 4 in ETC?
A: Cytochrome oxidase