Agbas (OxPhos)

Question 1

Ultrastructure of Mitochondria

Answer 1

• Two membranes: Outer (permeable: porin) and Inner (impermeable, metabolite transporters, cristae)
• Two compartments: intermembrane space and matrix (TCA Cycle and FA Oxidation)
• OXIDATIVE PHOSPHORYLATION occurs in INNER MEMBRANE

Question 2

Where does Oxidative Phosphorylation occur?

Answer 2

Inner Membrane of the mitochondria

Question 3

Oxidative Phosphorylation Overview

Answer 3

• TCA generates NADH and FADH2 –> high energy electrons flow through Electron Transport Chain (ETC)
• ATP synthase allow protons to return to matrix = ATP synthesis

Question 4

Successful Oxidative Phosphorylation must accomplish what 3 things?

Answer 4

1. transfer electrons from NADH/FADH2 to O2
2. establish proton gradient across inner mitochondrial membrane
3. synthesize ATP

Question 5

Electron Transport Chain Components

Answer 5

Respirasome (Complex I, III, IV) –> pump protons

1) NADH Q oxidoreductase
3) Q cytochrome c oxidoreductase
4) cytochrome c oxidase

• Complex II: Succinate Q reductase; uses FADH2
• Complex II does NOT pump protons

Question 6

Mobile Electon Carriers (2)

Answer 6

1. Coenzyme Q (ubiquinone)
   - transfers electrons from complex I/II to complex III
2. Cytochrome C
   - shuttles electrons from complex III to complex IV
   - final component of ETC
   - catalyzes reduction of O2

Question 7

Complex I

Answer 7

• NADH dehydrogenase or NADH-Q oxidoreductase
• first point of entry of electrons from NADH
• iron sulfur clusters: role in reduction rxns, proteins do NOT give up protons

Question 8

Complex II

Answer 8

• Succinate-Q reducatase
• FADH2 enters ETC (Connects TCA to Oxphos)
• does not pump protons, less ATP synthesized from oxidation of FADH2

Question 9

Complex III

Answer 9

• Q cytochrome C oxidoreductase
• passes electrons from QH2 to cytochrome C
• flow of electrons leads to transport of 2 protons to cytoplasmic side

Question 10

Complex IV

Answer 10

• Cytochrome C oxidase
• catalyzes transfer of electrons from reduced Cyt C to molecular oxygen, the FINAL ACCEPTOR
• makes reactions aerobic, makes humans breathe
• 4 electrons funneled to oxygen to reduce it to water
• 2 heme and 3 copper

Question 11

Free Radicals

Answer 11

• partial reduction of O2 generates high reactive oxygen derivatives, called reactive oxygen species (ROS)
• superoxide ion, peroxide ion, hydroxyl radical
• Normal Production: signaling (growth, hormone synth, inflammation)
• Over Production: damage to DNA, proteins, lipids

Question 12

Antioxidants (2)

Answer 12

1. Superoxide Dismutase (SOD) –> forms O2 and H2O2
2. Catalase –> H2O2 to O2 and 2 H2O

EX: glutathione peroxidase, Vit-E, Vit-C

Question 13

Mechanism of Action of SOD

Answer 13

1. SOD1: Cu/Zn SOD –> cytosolic

2. SOD2: Mn/Zn SOD –> mitochondrial

Question 14

Proton Motive Force

Answer 14

• two factors constitute a proton motive force to drive ATP synthesis by Complex V

1. pH gradient
2. Membrane potential

Question 15

Chemiosmotic Hypothesis

Answer 15

1. ETC moves protons across inner membrane as electrons flow from one complex to the next
2. ATP synthase uses proton motive force to phosphorylate ADP
3. If membrane disrupts, proton motive force cannot be established, and ATP synthesis does NOT occur

Question 16

ATP Synthase

Answer 16

• in inner membrane (ball and stick structure)
• F0 subunit: stick, has proton channel
• F1 subunit: ball (into matrix), catalytic domains
• only B subunit of F1 is catalytically active
• a/B subunits arranged in hexameric ring (y and epsilon above as stalk)
• form dimers that form oligomers (stabilize molecules to rotational forces of catalysis)
• MAINTAIN CURVATURE OF INNER MEMBRANE

Question 17

Cristae

Answer 17

allow the proton gradient to be in close proximity to the ATP synthase

Question 18

ATP Synthesis

Answer 18

• 1 mol of ATP requires 3 + 1 H passage

- oligomycin disrupts proton transport through the channel

Question 19

ATP-ADP translocase

Answer 19

• in outer/inner mito-membranes, works with mitochondrial carriers
• Flow of ATP and ADP coupled (ADP enters matrix only if ATP leaves)
• COMPLEX VI

Question 20

NADH2 Shuttle Systems

Answer 20

1. Malate-Aspartate Shuttle - oxaloacetate –> malate –> a-ketoglutarate (w/aspartate)
   - COMPLEX 1
2. Glycerophosphate Shuttle - G3P –> DHAP
   - CoQ

Question 21

Malate-Aspartate Shuttle

Answer 21

• heart, liver, kidneys
• generates NADH in mito-matrix
• NADH enters ETC at COMPLEX 1

Question 22

Glycerophosphate Shuttle

Answer 22

• skeletal muscle, brain
• generates FADH2 in inner mito-membrane
• FADH2 joins ETC at CoQ

Question 23

Inhibition of OxPhos

Answer 23

• when transfer of electrons inhibited = dec. in proton pumping, proton gradient and inhibition of ATP synth

Question 24

Uncoupling and Heat Generation

Answer 24

• used to generate heat and maintain body temperature (uncouple OxPhos from ATP Synth)
• in brown adipose tissue
• UCP1: thermogenin (uncoupling protein) –> transfer protons from cytoplasm to matrix side (heat)
• UCP2/UCP3: energy homeostasis

Question 25

Complex I inhibitors (4)

Answer 25

- amytal - rotenone - myxothiazol - piericidin A

Question 26

Complex II inhibitor

Answer 26

- malonate

Question 27

Complex III inhibitor

Answer 27

- antimycin

Question 28

Complex IV inhibitor

Answer 28

- CO, cyanide, H2S

Question 29

Complex V inhibitor

Answer 29

- oligomycin