Lectures 20-22 - Mitochondria and Oxidative Metabolism

Question 1

1) Define catabolism
2) What does it generate?
3) What mols does it involve?
4) Define Anabolism
5) What does it use?
6) What processes does it involve?
7) What is the relationship btw catabolism and anabolism?
8) What general process are these two a part of?

Answer 1

1) breaking down of mols
2) produces ATP (energy), NADH and NADPH (reduced) and generates CO2, H20 and NH3 as byproducts
3) Energy production, carbs, proteins, lipids
4) building of mols
5) uses ATP–> ADP + Pi; NADH –> NAD+ + H+; NADPH –> NADP+ + H+ (oxidized); produces heat
6) synthesis of macromolecules, muscle contraction, active ion transport, thermogenesis
7) rate of catabolism = rate of anabolism
8) metabolism

Question 2

1) Why can ATP be used for energy?
2) What is the rxn?
3) When can it be used?
4) What is required to regenerate ATP?

Answer 2

1) has 2 high E phosphate bonds that can be liberated through breaking the bond
2) ATP –> ADP + Pi; ATP –> AMP + 2Pi; hydrolysis used to cleave bond and break off P gp, thereby releasing energy
3) in very E favorable rxns
4) requires high E bonds to regenerate ATP

Question 3

1) What mols are used in catabolism to regenerate ATP, NADH..etc
2) What happens to these mols?
3) True or False: All Cs are created equally
4) Which mol has less E?

Answer 3

1) carbs, lipids
2) oxidized carbs are reduced

reduced lipids are oxidized

3) False –> more OH’s or Os attached to C, the more its oxidized –> less potential E that can fuel E production –> less Hs that can be extracted
4) carbs; lipids are typically always reduced (all branches used) –> more Hs available for extraction

Question 4

1) What are extracted Hs?
2) What is NADH made up of?
3) What part of the mol is used in redox reactions?
4) Which form is oxidized and which is reduced?
5) How is NADP look different?
6) What rxns are these mols used in?

Answer 4

1) NADH and NADPH
2) AMP mol + nicotinamide held together by phosphodiester bond
3) nicotinamide part
4) NADH (reduced)

NAD+ (oxidized)

5) hydroxyl gp on ribose sugar can be phosphorylated
6) NAD form used in redox rxns of catabolism

NADP used in rxns of anabolism

Question 5

1) Describe relationship with redox rxns and metabolism processes

Answer 5

1) reduced substrate –> oxidized in catabolism rxns –> oxidized product (C atoms in lipid mol are oxidized)

nucleotide captures e- released from oxidized product (NADP+ –> e- –> NADPH (reduced)

2) oxidized precursor –> reduced in anabolism rxns –> reduced product

nucleotide gives off e- to reduced product

(NADPH –> e- –> NADP+ (oxidized)

Question 6

1) What enzyme involves transfer of P gp (ATP –> ADP + Pi)
2) True or False: These rxns using nucleotides can either generate or utilize ATP
3) Give some examples

Answer 6

1) kinases
2) True
3) glu –> glu-6-P

Question 7

1) True or False: ATP is the only nucleotide with high E P bonds
2) How can NDP be built back up?

Answer 7

1) False; NTP –> NDP + Pi
2) Use P from ATP to produce NTP from NDP; nuceoside diphosphate kinase –> transfers P arounds btw nucleotides

NMP–>NDP from ATP –> transfers P btw ribonucleotides

Question 8

1) What is the main purpose of using Acetyl CoA?
2) describe general structure of acetyl CoA
3) Where can Acetyl CoA come from?

Answer 8

1) shuttling around this 2-C mol (acetyl gp)
2) ADP+pantothenic acid+S+acetyl gp (gp attaches to S atom)
3) carb (glycogen–>glucose–>pyruvate–>)

fat (triglyceride–> free FA –>)

protein metabolism (protein–>AA–>)

***can derive E from all major classes if convert to acetyl CoA

Question 9

1) What is Acetyl CoA used for/fed into?
2) Glucose –> pyruvate; What can pyruvate be used for?

Answer 9

1) TCA (tricarboxylic acid cycle)

ketone bodies –> from FA (biosynthetic rxn)

sterols and long chain FA

2) 3-C mol –> transamination –>alanine

carboxylation –>oxaloacetate

oxidative decarboxylation (loses e) –> acetyl CoA

reduction –> lactate

Question 10

Describe general pathway of pyruvate –> acetyl gp

pyruvate dehydrogenase PDP

Answer 10

1) acetyl gp w/ carboxyl gp attached

at end of rxn, acetyl gp will be transferred to Coenzyme A by sulfur to produce acetyl CoA

nucleotides that use B vit

pyruvate is oxidize –> acetyl gp carried to Coenzyme A and e- carried to NADH

connection btw carb metabolism and mitochondrial TCA cycle

Question 11

1) How is PDH regulated?
2) What is feedback inhibition
3) What does regulation depend on?
4) How does phosphorylation regulate?

Answer 11

1) feedback inhibition and by phosphorylation
2) products inhibit substrate binding site
3) rate it performs depends on [] of substrates

if prdt [] are building up, they can go back and inhibit rxn

4) phosphorylation inactivates PDH; desphosphorylation activates PDH

if prdt [] builds up –> activate kinase –> phosphorylate –> inactive

prdt [] low –> activate phosphatase –> dephosphorylate –> active

Question 12

1) What is common use of acetyl, CoA?
2) What are prdts/rxns?

Answer 12

1) shuttle it into TCA cycle

transfer of acetyl gp

2) 2 Cs leave as CO2

4 oxid-red rxns : 2 e-

1 GTP

eventually ~11 ATP produced by conservation of P bonds in ETC

Question 13

Describe TCA cycle

• Redox rxns
• ATP use

Answer 13

2 Cs from acetyl gp stay in mol throughout entire cycel

1) oxaloacetate + acetyl CoA –> citrate
2) citrate –> isocitrate
3) NAD+ –> NADH

product of dehydrogenation rxn –> CO2 and alpha ketoglutarate

4) NAD+ –> NADH, CO2

alpha ketoglutarate –> succinyl CoA

5) Succinyl CoA –> succinate

GDP –> GTP

6) Succinate –>Fumarate

FAD –> FADH2

7) Fumarate –> Malate
8) Malate –> oxaloacetate

NAD+ –> NADH

Question 14

1) How is TCA cycle regulated?
2) How does IsoC DH regulate?
3) How does a-KG DH regulate?

***also senses [] of GTP

Answer 14

1) feedback inhibition
- controlling PDH

(lots of Acetyl CoA and NADH - turn off)

2) Isocitrate DH –> 1st pair of NADH mols

senses ATP/ADP ratio

lots of ATP –> turns off –> dec activity

lots of ADP –> turns on –> inc activity

3) lots of NADH –> turn off –> dec activity

lots of NAD+ –> turn on –> dec activity

Question 15

1) What are other uses of TCA cycle?
6) What is the overall effect?

***if use intermediates, must regenerate them in other pathways to continue cycle

Answer 15

1) oxaloacetate –> AA synthesis
2) citrate –> FA and sterol synthesis
3) a-KG –> AA synthesis –> neurotransmitters
4) succinyl CoA –> heme synthesis

***can also be used to make ketone bodies

5) malate –> gluconeogenesis
6) effect of removing intermediates and regenerating OAA

Question 16

1) How does pyruvate –> OAA?
2) If acetyl CoA [] inc, what happens?
3) What does this rxn require?

Answer 16

1) pyruvate + HCO3- –>pyruvate carboxylase –> OAA
2) increases production of OAA; acetyl CoA - positive allosteric effector (won’t make OAA unless there’s enough acetyl CoA
3) energy, ATP

***biotin used in carboxylation rxn

Question 17

1) How many membranes in a mitochondria?
2) what is the key permeability barrier
3) Where is ATP synthase/ETC located?

Answer 17

1) inner and outer
2) inner mitochondrial membrane
3) folds of inner membrane

Question 18

1) What enters Complex I
2) What enters Complex II
3) What does UQ do?
4) What does cytochrome C do?
5) What happens in Complex IV
6) True or False : ETC is just a series of chemical ox-red rxns

Answer 18

1) e- that were passed from NAD+ –> NADH enter here
2) e- from FADH2 enter here
3) ubiquitone transfers e- from complex I and II to complex III

UQ –> UQH2

4) connects complex III and IV
5) draws e- towards O2 –> reduced to form H20
6) True; e- passed from one carrier to next; given to a carrier with a higher affinity –> O2 has greatest affinity of all

Question 19

1) Describe complex I

***e- go into 4 diff e- acquiring mols

7) What is the E conserving step?

Answer 19

1) NADH drops off 2 e-
2) FMN (flavinmononucleotide) picks them up
3) FMN –> FeS (iron-sulfide containing protein); can only carry 1 e- at a time
4) FeS –> FeS, 1 e-
5) FeS –> N-2, 1 e-
6) N-2 –> UQ –> UQH2 (reduced), 2 e-
7) pumps protons from matrix side into intermembrane space –> creating [] gradient

Question 20

1) Describe complex II
3) What is main difference btw this complex and complex I
4) Describe complex IV
5) How many e- are required to reduce O2?

Answer 20

1) e- from FAD enter complex II –> transfered to UQ
2) FA oxidation produces FADH2
3) no proton pumping
4) cyt c brings e- into complex –> mediated by heme rings with Fe in center (1 e- at a time)
5) 4e- required to reduce O2 into 2H20; 4 H+ pumped into intermembrane space

Question 21

1) Which complexes pump protons?
2) What substrates generate NADH?
3) What substrates generate FAD?
4) What are inhibitors of Complex I?
5) What are inhibitors of Complex IV?

Answer 21

1) I, III and IV
2) malate, a-KG, isocitrate

FA and ketone oxidation

3) succinate

Fatty Acyl CoA

a-glycerol P

4) rotanone
5) CO, KCN, potassium azide

***inhibit proton pumping

Question 22

1) What is main goal of ETC?
2) What produces ATP?
3) Describe process

Answer 22

1) creating proton and charge gradient –> inc pH of intermembrane space; lots of + outside, lots of - inside
2) ATP synthase –> extends into membrane; end of it extends into matrix
3) selective re-entry of protons back into matrix (from intermembrane side)

H+ goes down [] gradient

transfer P to ADP –> ATP

Question 23

1) What is the big picture of ATP/energy?

Answer 23

1) balance btw E production and E utilization
2) faster you use E, the more you have to make E

Question 24

1) What does this diagram show?
2) What happens when you add substrate from TCA cycle (succinate)
3) What if inc ADP []?

Answer 24

1) The rate at which the mitochondria are respiring by measuring O2
2) not much, little decrease in O2, but no stimulus
3) inc in ADP –> must make ATP –> drop in O2 [] (use in ETC)

eventually all ADP is converted to ATP, so returns back to baseline [] of O2 (rate slows)

Question 25

1) How do substrates and ATP get into mitochondria?

give exs

2) What kind of transporter is ADP/ATP?
3) What kind of transporter is P/H+?
4) Are both required for ATP synthesis to occur?

Answer 25

1) transporters

phosphate –> phosphate transporter (coupled with H+ coming in to set up [] gradient)

ADP –> adenonie nucleotide translocase

***ATP brought out to cytosol to be used and then ADP brought in to mitochondria to be phosphorylated

2) antiporter (ADP in, ATP out)

more + outside, helps drive more - charge ATP out and bring ADP in

3) symporter (both coming in)

protons going into mitochondria, coupled to bring P in as well

4) YES!

Question 26

1) Does citrate wait around for TCA cycle?
2) What is ETC coupled with?
3) Can ETC be uncoupled?
4) Explain
5) What’s its effect on ATP synthesis, etc

Answer 26

1) no, under certain circumstances it will eave mitochondria for FA and sterol biosynthesis

coupled with Malate in antiporter (malate in, citrate out)

2) rate of ATP synthesis (based on [] of ATP/ADP
3) yes, 2,4-dinitrophenol (proton ionophore that equilibriates pH across the inner mitochondrial membrane
4) On phenol, theres a dissociative ion

dinitrophenol goes from outside mitochondrial space (low pH) into matrix

proton dissociates off –> charged ion, weak acid, lipid soluble

5) carries a proton back into matrix and bypasses ATP synthase therefore creates an alternate path for getting protons back into matrix –> can’t use H+ for ATP synthesis –> destroys proton gradient

Question 27

1) What does diagram show?
2) Describe ex with brown adipose tissue in young animals
3) what is the result?
4) What activates this process?

Answer 27

1) add dinitrophenol –> brings protons back into matrix –> inc proton pumping and ETC –> using more O2 and results in steeper rate of O2 use
2) UCP 1 - uncoupling protein 1 –> allowing proton movement back into matrix without going through ATP synthase
3) all redox rxns used to make proton gradient is wasted E –> converted into heat –> E is not conserved –> not conserving ATP bonds
5) cold sensor in hypothalamus –> norepinephrine released –> G protein complex –> PKA

Question 28

1) Features of Mitochondrial genome
5) True or False: mitochondria are maternally inherited
6) What are some effects of mutations in genes controlling mitochondrial fn?

Answer 28

1) genes for 13 proteins
2) components of complex I, III, IV as well as ATP
3) several rRNAs and tRNAs
4) 90+ % of their proteins come from nuclear genes in nucleus –> difficult to get proteins translated in cytoplasm into mitochondria
5) True
6) dec ETC and ATP synthase activity

build up of lipids and other mito substrates

neuro deficiencies (1st affected) - neurons rely on Na+/K+ ATPase (need ATP)

muscle weakness and exercise intolerance

Question 29

1) What are the reactive O2 intermediates
2) what happens when there’s incomplete reduction of O2 in ETC?
3) Describe rxns

Answer 29

1) Superoxide, O2-

hydrogen peroxide H2O2

hydroxyl radical OH*

2) reactive O2 intermediates are created
3) O2 + 1e- –> O2-

O2- + 1e- –> H2O2

H2O2 + 1e- + H+ –> OH*

OH* + 1e- –> H20

***all can cause serious damage

Question 30

1) Describe natural formation of reactive O2 intermediates
2) What can happen with UQ?

Answer 30

1) Fe+2 + H2O2 –> Fe+3 + OH* + OH-

O2- + H2O2 –> + H+ –> O2 + H2O + OH*

2) It can be leaky –> O2 can come in and steal an e- and become superoxide O2-

Question 31

1) What are the consequences of reactive O2 species

Answer 31

1) Lipid Peroxidation

• unsaturated FA in phospholipids –> extraction of a H atom from a dbl bond by an OH –> alters fluidity of membranes
• lipid radicals react with O2 to form lipid peroxidase
• aging spots –> build ulp
• leaky membranes –> change poracity in membrane

2) Protein oxidation
* attack AA side chains –> must be degraded by intracellular proteases
3) DNA mutations –> particularly in mito DNA –> more free radicals

Question 32

1) What’s one reason reactive O2 intermediates are generated naturally?
2) describe
3) How are reactive O2 intermediates removed?

Answer 32

1) destroy bacteria
2) O2 –> O2- –> H2O2 –> OH* (in phagocyte)

bacteria are engulfed into phagocyte –> react with radicals and are destroyed

3) 2O2- –> superoxide dismutase (2H+ enter) –> one molecule becomes O2 and other H2O2 –> catalase (H2O2 enters ) –> 2H2O + O2

H2O2 reacts with Fe+2 to create OH*

***nothing to combat OH* so will go do damage