Bioenergetics + resp. chain

Question 1

When can a reaction considered to be exergonic or endergonic?

Answer 1

depends on ΔG = change in (useful) Gibb’s free energy

• ΔG negative → E is released during RXN, happens spont., irreversible + exergonic
• ΔG = 0 → RXN is in equilibrium, reversible
• ΔG positive → E is consumed, does not happen spont., endergonic

Question 2

Metabolism is the combination of anabolic and catabolic processes.

Which of those can be considered exergonic or endergonic?

Answer 2

anabolism = endergonic, E must be invested

catabolism = exergonic, E can be gained

Question 3

What are auto- and heterotrophic organisms?

Answer 3

• *heterotrophic:** couple metabolism to breakdown of complex organic molecules
• (lit. use others for nutrition)*
• *autotrophic:** gain E from producing complex organic molecules from simple substances (i.e. photosynthesis)
• (lit. self nourishing​)*

Question 4

Define group transfer potential.

Answer 4

ΔG that is released when high-energy bond is cleaved

Question 5

List some high-energy compounds.

Answer 5

• ATP: energy currency in body
• phosphagens: storage form of high-E phosphates
• thiol esters: i.e. acetyl-CoA, acyl carrier protein
• AA esters: involved in protein synthesis
• SAM = active Met: used for transfer of -CH₃
• UDPGl: involved in glycogenesis
• PRPP: involved in purine synthesis

_{rather overview than list to memorize}

Question 6

What are the most important sources of high-E phosphates?

Answer 6

• oxidative phosphorylation: in resp. chain
• glycolysis
• substrate-level phosphorylation: in TCA
• mito. PEPCK: part of gluconeogenesis
• through other phosphagens
• interconversion reactions

_{<strong>​</strong>rather overview than list to memorize}

Question 7

What are the classical phosphagens in vertebrates and invertebrates?

Answer 7

phosphagen = storage form of high-energy phosphate

• vertebrates = creatine phosphate
• invertebrates = Arg phosphate

Question 8

What is the difference btw substrate-level phosphorylation and oxidative phosphorylation?

Answer 8

• substrate-level phosphorylation: directly phosphorylating ADP with P_i, energy provided from a coupled reaction
• oxidative phosphorylation: ATP generated from the oxidation of NADH/FADH2 and the subsequent transfer of e^- and pumping of H⁺

Question 9

Succinyl CoA synthetase has a special role in the synthesis of high-energy phosphates.

Explain w/r/t its structure.

Answer 9

enzyme of TCA that catalyzes the only substrate-level phosphorylation, either ATP or GTP, dependent on tissue

succinyl-CoA + GDP/ADP + Pi ⇔
succinate + GTP/ATP + CoA-SH

has 2 subunits:

• α = G1, same in every enzyme
• β = G2 or A2 → phosphorylates either GDP/ADP

Question 10

In which tissues is GTP, in which ATP produced by succinyl CoA synthetase?

Answer 10

• in tissues that are mainly involved catabolism (heart, brain)
  → A2 subunit, hence synthesizing ATP
• in tissues that are mainly involved in anabolism (liver, kidney)
  → G2 subunit, hence synthesizing GTP

Question 11

What are interconversion reactions?

List 2 important enzymes catalyzing such interconversions.

Answer 11

conversion of mono-/di-trinucleotides into each other

• adenylyl kinase
• nucleoside diphosphate kinase

Question 12

Which reaction is catalyzed by adenylyl kinase?

Answer 12

ATP + AMP ⇔ 2 ADP​

Question 13

Which reaction is catalyzed by nucleoside diphosphate kinase?

Answer 13

converts nucleotides,

e.g. ATP + UDP ⇔ ADP + UTP

Question 14

Which enzyme ensures the irreversibility of reactions where ATP is cleaved to AMP?

(e.g. acyl-CoA synthetase in the synthesis of long-chain FAs)

Answer 14

inorganic phosphatase

PP_i + H₂O → 2P_i

Question 15

List important electron carriers.

Answer 15

• NAD⁺, NADP⁺: derivatives of vitamin niacin
• FAD⁺, FMN: derivatives of vitamin riboflavin
• ubiquinone: involved in resp. chain
• Fe-S complexes: involved in resp. chain
• heme: enzyme-bound prosthetic group

_{again, rather overview than list to memorize​}

Question 16

How does heme transfer electrons?

Answer 16

strictly speaking it only transfers 1 e^- at a time

by oxidation of Fe²⁺ to Fe³⁺

NOTE: although Hb, and myoglobin have heme center, those are NOT oxidized (physiologically)

Question 17

What are oxidoreductases?

Classify into 4 groups.

Answer 17

enzymes involved in oxidation/reduction

• oxidases
• dehydrogenases
• hydroperoxidases
• oxygenases

Question 18

Which reaction is catalyzed by oxidases?

Answer 18

removal of H₂ from substrate, using oxygen as acceptor to form water/hydrogenperoxide

AH₂ + 1/2 O₂ → A + H₂O/H₂O₂

Question 19

List 3 important oxidases.

Answer 19

• cytochrome c oxidase (complex IV): terminal component in resp. chain
• flavoproteins: enzymes containing FAD/FMN as prosthetic group
• metalloflavoproteins: enzymes that use metal as cofactor AND contain FAD/FMN as prosthetic group

Question 20

What are the 2 functions of dehydrogenases?

Which reaction do they catalyze?

Answer 20

• transfer H₂ from substrate to another (basically same as oxidases, but DO NOT use oxygen as acceptor)
  AH₂ + carrier → carrier-H₂ + A
  → often use NAD/NADP/FAD/FMN as carrier
• transfer e^- in resp. chain

Question 21

Which dehydrogenases are NAD- which are NADP-linked?

Answer 21

• NAD-linked dehydrogenases catalyze reactions of oxidative pathways (glycolysis, TCA, resp. chain
• NADP-linked dehydrogenases catalyze reactions of reductive snythesis (lipogenesis, steroid synthesis, PPP)

Question 22

What are the 2 groups of hydroperoxidases?

Which reactions do they catalyze?

Answer 22

​both reduce H₂O₂, substrates are different though

• peroxidases: use additional e^- acceptor
• *H₂O₂ + AH₂ → 2 H₂O + A**
• catalases: use H₂O₂ as donor and acceptor
• *2** H₂O₂ → 2 H₂O + O₂

​REMEMBER: catalases are one of the fastest enzymes

Question 23

What are oxygenases?

Differentiate.

Answer 23

transfer and incorporate oxygen into substrate, in 2 steps

1. O binds to enzyme at active site
2. bound O reduced/transferred to substrate

→ mono-/dioxygenases

Question 24

Which reactions are catalyzed by mono- and dioxygenases?

Answer 24

• monooxygenases incorporate 1 O atom, other O reduced to water
• *AH + O₂ + ZH₂ → A-OH + H₂O + Z**
• dioxygenases incorporate both O into substrate
• *A + O₂ → A-O₂**

Question 25

Cytochromes can also be regarded as... which group of enzymes? In which organelles can they be found?

Answer 25

**dehydrogenases** (_except_ cytochrome c oxidase obv) * in mitochondria (cytochrome b, c1, c) * in ER (CYP, 450 b5) _NOTE:_ **CYP450 rather monooxygenases** than dehydrogenases

Question 26

To which group of enzymes does superoxide dismutase belong? Which reaction does it catalyze?

Answer 26

**= monooxgygenase** protects aerobic organisms from ROS **O₂^- + O₂^- + 2H⁺ → H₂O₂ + O₂**

Question 27

Superoxide dismutase has 2 isoenzymes. Where can they be found?

Answer 27

* **SOD1** in cytosol * **SOD2** in mitochondria

Question 28

Which structure are involved in the resp. chain?

Answer 28

* uses 3 complexes (**I, III, IV**) for transport of ^- from _NADH/H⁺_ * uses 1 complex (**II**) for transport of e^- from _FADH₂_ via **CoQ** and **cytochrome c₁** to reduce O₂, forming H₂O _meanwhile:_ **H⁺ pumped** into IM space, later used for **ATP synthesis**

Question 29

What happens at complex I of the resp. chain? Name.

Answer 29

**_NADH dehydrogenase_** 1. **NADH/H⁺ is oxidized** to NAD⁺ 2. its 2e^- are transferred via _FMN_, and a series of _Fe-S centers_ to 3. reduce CoQ → **ubiquinol** (QH₂) _additionally:_ **4H⁺ are pumped** from matrix into IM space

Question 30

What is the function of complex II? Name. Which reaction does it catalyze?

Answer 30

**_succinate dehydrogenase_**​ **​succinate + FADH₂ → fumarate + FAD⁺** * also participates in _TCA_ * 2e^- from FADH₂ also transferred via _Fe-S centers_ to reduce CoQ → **ubiquinol** (QH₂) _NOTE:_ does NOT pump any H⁺ into IM space

Question 31

What are the 3 forms of ubiquinone? Where is it located?

Answer 31

in _inner mitochondrial membrane_ * **ubiquinone** (= CoQ): fully oxidized * **semiubiquinone** (= QH): partially reduced * **ubiquinol** (= QH₂): fully reduced

Question 32

What is the function of CoQ?

Answer 32

is **reduced by complex I _or_ II**, then diffuses in inner mitochondrial membrane **to complex III** to _transfer its electrons_

Question 33

What happens at complex III of the resp. chain? Name.

Answer 33

**_cytochrome c dehydrogenase_** _**Q cycle** happens:_ 1. 2e^- from ubiquinol (QH₂) transferred via _cytochrome b_ and _Rieske Fe-S center_ 2. to **cytochrome c₁** (now CoQ reformed) _NOTE:_ here also **4 H⁺ pumped** from matrix into IM space

Question 34

What is the function of cytochrome c1? Where is it located?

Answer 34

**reduced by complex III**, then diffuses in IM space (NOT in inner mitochondrial membrane) to **complex IV** to _transfer its electrons_

Question 35

What happens at complex IV? Name.

Answer 35

**_cytochrome oxidase_** 1. cytochrome c₁ oxidized to transfer its 2e^- via _2 heme groups_ and _2 Cu centers_​ to complex IV 2. catalyzes: * *1/2 O₂ + 2H⁺ → H₂O** ​_NOTE:_ unlike complex I and III, complex IV **pumps _only 2 H⁺_** from matrix into IM space

Question 36

What does the chemiosmotic theory state?

Answer 36

**movement of H⁺ across inner mitochondrial membrane drives ATP synthesis** due to electrochemical gradient established by _complexes I, II, IV_ * complex I, III pumping 4H⁺ into IM space (each) * complex IV pumping 2H⁺ into IM space

Question 37

Which enzyme eventually catalyzes the synthesis of ATP? Describe its function w/r/t its structure.

Answer 37

**_ATP synthase_** _has 2 subunits:_ (dimer...?) * **F₀ complex** (in _membrane_): acts as _H⁺ channel_, H⁺ diffuse from IM space into matrix * **F₁** **complex** (in _matrix_): _phosphorylates_ ADP to ATP ​→ phosphorylates 1 ATP/4H⁺ | (F-type ATPase)

Question 38

What is the P/O ratio of oxidative phosphorylation?

Answer 38

**ratio of ATP produced per oxygen _atom_ reduced by resp. chain** * _for NADH-linked substrates:_ **2.5** b/c 10H⁺ pumped into IM space per NADH * _for FADH-linked substrates:_ **1.5** b/c only 6H⁺ pumped into IM space per FADH₂

Question 39

Which complex is blocked by TTFA? What is another inhibitor that acting on the same target?

Answer 39

inhibits **complex II** also: **malonate** = competitive inhibitor

Question 40

Which complex is blocked by barbiturates like amobarbital? Which other substances do you know that have the same effect?

Answer 40

inhibit **complex I** also: **piericidin A, rotenone** (insecticide)

Question 41

Which complex is inhibited by the antibiotic antimycin A? Again, name another drug that does the same.

Answer 41

**complex III** also: **dimercaprol** (chelation therapy in metal toxicity)

Question 42

Which complex is blocked by CO? Give other examples having the same effect.

Answer 42

**complex IV** also: **H₂S, CN^-**

Question 43

Think of the lab exam... What do atractyloside and oligomycin do?

Answer 43

both inhibit ATP synthesis via different mechanisms * **atractyloside:** inhibitor of _ATP:ADP translocase_ * **oligomycin:** blocks F₀ complex of _ATP synthase_

Question 44

What are uncouplers? Give 2 examples.

Answer 44

**dissipate H**⁺ **gradient** via incr. the permeability of the inner mitochondrial membrane = _H⁺ ionophores_ → E used for generation heat instead of driving ATP synthase * **2,4-dinitrophenol:** classic uncoupler * **thermogenin:** in brown adipose tissue _NOTE:_ also support chemiosmotic theory

Question 45

What is respiratory control?

Answer 45

oxidative phosphorylation is (physiologically) **regulated by the amount of ADP** that can be used for ATP synthesis

Question 46

NADH is gained during cytosolic glycolysis can also be used to fuel oxidative phosphorylation. But How does it get into the mitochondrial matrix?

Answer 46

* **malate-Asp shuttle** forms intermediates, eventually NADH resynthesized _in matrix_ * **glycerophosphate shuttle** converts NADH to FADH₂ in _inner mitochondrial membrane_

Question 47

Describe the mechanism of the malate-Asp shuttle.

Answer 47

1. **_malate dehydrogenase_** in IM space/cytosol OXA + NADH → malate + NAD⁺ **_ASAT_** in matrix OXA + Glu → α-KG + Asp 2. **2 transporters** shuttle malate and Asp across the inner mitochondrial membrane * _OGC:_ malate - α-KG antiporter * _AGC:_ Asp - Glu antiporter ⇒ **same enzymes** then catalyze reverse reactions in the other compartment → NADH can then be used for resp. chain

Question 48

Explain the mechanism of the the glycerophosphate shuttle.

Answer 48

1. **_cytosolic glycerol-3P dehydrogenase_** NADH + DHAP → NAD + G3P 2. G3P crosses outer mitochondrial membrane 3. **_mitochondrial glycerol-3P dehydrogenase_** 1. **_​_**G3P + FAD → DHAP + FADH₂ 2. FADH₂ then transfers its 2e^- to _CoQ_ 4. DHAP returns into cytosol