Electron Transport & Oxidative Phosphorylation Flashcards by Ann M

What is notable about the outer mitochondrial membrane?

Permeable
Porins for ion transport
Transport proteins for larger molecules

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What is notable about the inner mitochondrial membrane?

Only permeable to O2, H2O, CO2
Rich in proteins (incl. ETS)
Cardiolipin-rich (forms cristae)
Intercristal spaces with high [H+]

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What is contained within the mitochondrial matrix?

Soluble enzymes of oxidative metabolism
Substrates
Cofactors
DNA & RNA
Ions

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What are the two ways that NADH’s electrons are transferred into the matrix?

Malate-aspartate shuttle
Glycerophosphate shuttle

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Where is the malate-aspartate shuttle found?

Heart, liver, kidneys

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Where is the glycerophosphate shuttle found?

Skeletal muscle

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How does the malate-aspartate shuttle bring OAA into the mitochondrion?

Oxaloacetate reduced back to malate
Malate brought into the mitochondrion
Malate oxidized back to OAAH (producing NADH)

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How does the malate-aspartate shuttle get OAA out of the mitochondrion?

An amino group from glutamate is transferred over to OAA
Formed aspartate and a-ketoglutarate are transferred out to the cytosol, then converted back to OAA and glutamate (producing NAD+)

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Broadly speaking, how does the glycerophosphate shuttle work?

It transfers cytosolic NADH’s electrons over to mitochondrial FADH2 for entry into the ETS.

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What is the first reaction of the glycerophosphate shuttle?

3-phosphoglycerol dehydrogenase oxidizes NADH back to NAD+, which re-enters glycolysis (enzyme has electrons).

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What is the second reaction of the glycerophosphate shuttle?

DHAP is converted to phosphoglycerol.

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What is the third reaction of the glycerophosphate shuttle?

Flavoprotein dehydrogenase (in the IMM) gives phosphoglycerol’s electrons to FAD.

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What molecule carries electrons between complexes I, II, and III?

CoQ/ubiquinone

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What molecule carries electrons between complexes III and IV?

Cytochrome C

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Complex I enzyme

NADH-Coenzyme Q Oxidoreductase

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What is notable about the structure of complex I?

Large
L-shaped
Fe-S clusters
Proton wires

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What are the prosthetic groups on Complex I?

FMN and Fe-S clusters

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What is electron flow through Complex I?

NADH –> FMN –> Fe-S –> CoQ

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How many protons does Complex I pump into the matrix? When does proton movement occur?

4 protons
Moved as electrons pass through Fe-S clusters

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What drives electron transport in complex I?

Conformational changes that alter side-chain pK values, causing them to accept/release protons.

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Proton Wires

Arrangements of H-bonded groups in complex I that provide a way for protons to enter the IMS

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Complex II enzyme

Succinate CoQ oxidoreductase

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What is notable about Complex II?

Also participates in TCA cycle
Does not pump protons
Mushroom-shaped (C&D components in IMM)
Bound FAD

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How many Fe-S clusters does Complex II have?

3 Fe-S clusters

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How many Fe-S clusters does Complex I have?

8 Fe-S clusters

What is the movement of protons through complex II?

FAD/FADH2 --> Fe-S clusters --> CoQ

Cytochromes

Redox-active proteins that contain heme groups that cycle through Fe(II) and Fe(III) oxidation states.

Complex III enzyme

CoQ-Cytochrome C Oxidoreductase

What is notable about Complex III?

- Home to Q cycle - Rieske Fe-S cluster - Pear-shaped - 2 cyt Bs, 1 cyt C

What is different about a Rieske complex?

The Fe is coordinated by His rather than Cys

Pathway of the first electron in the Q cycle

CoQ --> Rieske --> Cyt C1 --> Cyt C --> Complex IV

Pahtway of the second electron in the Q cycle

CoQ --> Heme Bl --> Heme Bh --> Q (N site)

What happens to the two protons donated from ubiquinol in the Q cycle?

They enter the IMS

What are the primary outcomes of the Q cycle?

- 4 protons pumped into IMS - 2 taken up from matrix (to balance Q- charge) - 2 cytochrome Cs transported to CIV

Complex IV enzyme

Cytochrome C oxidase

What is notable about Complex IV?

- Contains the O2 - CuA/CuA center - Heme a/a3/CuB center

Pathway of first electron through Complex IV

Cyt C --> CuA/CuA center --> Heme A --> Heme A3 --> CuB

What happens to copper in complex IV?

It is reduced from Cu2+ to Cu+

Pathway of the second electron through Complex IV

Cyt C --> CuA/CuA --> Heme A --> Heme A3

What happens to the iron in Heme A3?

It is reduced from Fe3+ to Fe2+

What happens once Heme A3 and CuB are reduced?

- A peroxide bridge is formed between Heme A3 and CuB - 2 protons are extracted from the matrix, and used to break the bridge (Heme A3-OH and CuB-OH) - Two more protons are extracted from the matrix to form water and regenerate Heme A3 and CuB

How many protons are pumped into the IMS from Complex IV?

4 total (2 per pair of cytochrome Cs)

How many protons are pumped into the IMS from Complex III?

4 total (2 per Q cycle)

What is the net of the Complex IV reactions?

- 4 cytochrome Cs donating electrons - One O2 + 4 matrix protons forming 2 waters (1/2 O2 per cyt C pair)

Complex V enzyme

ATP synthase

Is complex V considered part of the ETS?

Chemiosmotic Theory

- Free energy of electron transport is conserved by creating an electrochemical gradient - Electrochemical gradient is harnessed to create ATP

What are the components of the Fo subunit of ATP synthase?

- C subunit ring - A subunit bound to the C subunit

What are the components of the F1 subunit of ATP synthase?

- 3 alpha-beta subunits - Gamma subunit stalk connecting to C subunits

What are the components of the protein stalk in ATP synthase?

- B subunit ("stator") - Delta subunit connecting the stator to F1

What are the three conformational states of the alpha-beta subunits on ATP synthase?

Open, loose, tight

Open conformation

Can't bind nucleotides (Beta E)

Loose conformation

Can bind ADP and Pi (Beta DP)

Tight Conformation

Can bind ATP (Beta TP)

How does the alpha beta subunit binding change mechanism work?

- ADP and Pi bind at the L site - Conformational change converts the L site to a T site, catalyzing ATP synthesis - ATP is synthesized at the T site as it is released from the O site on another subunit

What type of energy is associated with the electrochemical gradient?

Free energy

What type of energy is associated with ATP synthase?

Mechanical energy

What type of energy is associated with ATP?

Chemical energy

What is the structure of the C subunit

2 alpha helices connected via a loop turn

What 2 residues on the C subunit can pick up protons?

Glu or Asp

How does the C subunit rotate?

- Negative Glu/Asp picks up a proton - Change in hydrophobicity causes movement towards the IMM (rotation) - Proton enters a proton release channel and moves into the matrix

How many ATP are produced per turn of ATP synthase?

3 ATP total (1 ATP per alpha-beta subunit)

How many C subunits do humans have?

How many total protons are pumped into the IMS per glucose molecule?

112

What does the P/O ratio relate?

The amount of ATP synthesized to the amount of oxygen reduced

How is the P/O ratio calculated?

Protons pumped into IMS/Protons per ATP

How many protons are needed to produce 1 ATP?

Usually 4

What are two factors that change the P/O ratio experimentally?

- A proton is brough back into the matrix with pyruvate via the symporter (MPC) - ADP and ATP are exchanged at a 1:1 ratio, leading to a reduction in membrane potential

What are two phenomena that cause oxidative phosphorylation to be uncoupled from electron transport?

2, 4-DNP Brown adipose tissue

How does 2,4-DNP uncouple oxidative phosphorylation from electron transport?

- 2,4-DNP can cross the membrane since it's lipophilic and planar - It dissipates the gradient by grabbing protons and then diffusing back into the matrix - Dissipation of gradient causes release of thermal energy

How does BAT uncouple oxidative phosphorylation from electron transport?

- BAT contains UCP1 (thermogenin) in its IMMs - UCP1 is a symporter that transports anions and protons, but the anions can't dissociate from the protein. - Protons can reenter the matrix via UCP1, bypassing ATP synthase. - Energy released as non-shivering thermogenesis

What are the major structural components of NADH?

- Adenosine monophosphate (AMP) - Nicotinamide mononucleotide (NMN) - Nicotinamide (NAM)

Which part of NAD is capable of accepting the hydride?

The topmost carbon on the aromatic ring of the NAM

What are the major structural components of FAD?

- Riboflavin (ribitol & FAD) - Phosphoryl group - Adenosine