Electron transport Electron transport chainchain

Dr. SaidunnisaProfessor of Biochemistry

Matrix H+

+ NADH NAD+ + 2H+ 2H+

+ ½ O2 H2O 2 e

I Q III IV

Intermembrane Space cyt c

Learning objectivesLearning objectives At the end of the session the student

shall be able to define, describe, enumerate, analyze and list:

1. Location of ETC2. Components of ETC3. Sites of ATP production4. Oxidative phosphorylation5. Chemiosmotic theory 6. Energetics of ETC

I. Case:I. Case:A 35-year old male is rescued from

enclosed fire. Upon arrival in the emergency

department, he is disoriented and in moderate distress. He is coughing up soot, and has difficulty breathing.

Initial vital signs: BP 90/60, HR 120, RR 30, O2 sat 95%On examination: burned nasal hair, soot

around mouth, burns to face, arms and back.

What are the immediate concerns?

1Case conti…1Case conti…We have a tendency to focus on CO

as the diagnosis in victims of smoke inhalation.

Pitfall – CN exposure is frequently

overlooked. Consider that CN can be produced from the combustion of paper, silk, wool, plastic, and cotton.

The probability of CN exposure in fires is therefore high.

1. Case conti……1. Case conti……Lets us unfold the mystery of CO

and CN poisoning on human health while studying respiratory chain.

Stages of oxidation of food Stages of oxidation of food stuffsstuffs

First stage: Digestion in the GIT converts the

macro molecules into small units. (carbohydrates into glucose, lipids into fatty acids, proteins into amino acids).

This is called primary metabolism.

Introduction Introduction

Second stageSecond stage

These products are absorbed in mitochondrial citric acid cycle to generate reducing equivalents NADH or FADH2.

This is called secondary or intermediary metabolism.

Third stageThird stage

These reducing equivalents enter into the electron transport chain or respiratory chain where energy is released (ATP).

This is tertiary metabolism or internal or cellular respiration.

The energy (ATP) is then used for body synthetic purpose.

Cellular RespirationCellular Respiration

ATP-Universal currency of ATP-Universal currency of energy in living cellsenergy in living cells

The energy released from the hydrolysis of ATP is utilized for

Mechanical -muscle contractionTransport work -Sodium potassium

ATPase pumpBiochemical work -Initial steps of

GlycolysisAnabolic pathways -TAG, DNA, Protein

synthesisDetoxification(urea cycle),formation of

active intermediates like UDP glucose

HEAT

Chemistry of ATP hydrolysisChemistry of ATP hydrolysisThere are two high energy bonds in ATP

which are represented by a squiggle bond () on hydrolysis each releasing -7.3kcal/ mole.

Any bond that can be hydrolyzed with the release of same energy as ATP hydrolysis is called as “high energy bonds”.

More than 90% of ATP is formed through ETC.

Synthesis of ATP:Oxidative phosphorylationSubstrate level phosphorylation

High energy compoundsHigh energy compounds

High energy compounds:

ATP, GTP, UTP,PEPCarbamoyl phosphatecAMP 1-3

BisphosphoglycerateCreatine PhosphateAcetyl Co ASAM

Low energy compounds

AMPGlucose-1

phosphateFructose-1-

phosphate

Redox potentialsRedox potentialsOxidation: loss of

electronsReduction: gain of

electronsRedox couple: when

a substance exists both in the reduced and oxidized state.

Redox potentials: electron transfer potential E0’.

Substrate level Substrate level phosphorylationphosphorylationEnergy from a high energy

compound is directly transferred to ADP or GDP to form ATP or GTP without the help of electron transport chain.

Examples:1.Bisphosphoglycerate kinase

(glycolysis)2.Pyruvate kinase (glycolysis)3.Succinate thiokinase (TCA cycle)

Biological oxidationBiological oxidation

Definition: Transfer of electrons from reduced coenzymes through ETC to oxygen.

Energy released during this process is trapped as ATP.

This coupling of oxidation with phosphorylation is called oxidative phosphorylation.

In the body this is carried out by successive steps of dehydrogenations.

NADNAD+ + Linked Linked dehydrogenases dehydrogenases NAD+ is derived from nicotinic acid a

Vitamin B-complex. When NAD+ accepts the two hydrogen

atoms, one of the hydrogen atom is removed from the substrate as such the other is split into one hydrogen ion and one electron.

The electron is also accepted by the NAD+ so as to neutralize the charge on the co-enzyme.

H2 H + H+ + e-

AH2 + NAD+ A + NADH + H+

NADNAD+ + Linked Linked dehydrogenases dehydrogenases Examples: 1.Glyceraldehyde -3-phosphate

dehydrogenase.2.Isocitrate dehydrogenase3.Glutamate dehydrogenase4.Pyruvate dehydrogenase5.Alpha ketoglutarate

dehydrogenase

FADFAD Linked Linked dehydrogenases dehydrogenases FAD is derived from riboflavin a

Vitamin B-complex. Both the hydrogen atoms are

attached to the flavin ring. AH2 + FAD FADH2

Examples:1.Succinate dehydrogenase2.Fatty acyl CoA dehydrogenase.

Mitochondrial Mitochondrial OrganizationOrganization

Components of ETCComponents of ETC5 Complexes1. Enzyme complex I, (NADH

dehydrogenase)2. Enzyme complex II (Succinate

dehydrogenase)3. Enzyme complex (III)

Cytochrome reductase 4. Enzyme complex (IV)

Cytochrome oxidase. 5. ATP Synthase (V)

Components of ETCComponents of ETC

Two mobile carriers These are connected by two mobile

carriers Coenzyme Q, Cytochrome C.

Coenzyme Q, connects complex-1 and 11.

Cytochrome C connects complex 111 and 1V

Electrons flow from more electronegative to electropositive components.

Complex-IComplex-I

Complex- IVComplex- IV

Coenzyme Q

Iron sulfur proteins and Iron sulfur proteins and CytochromesCytochromesIron sulfur proteins: They exist in the

oxidized (Fe+3) or reduced (Fe+2) on accepting an electron.

Cytochromes: Cytochromes: Cytochrome C is a mobile component of ETC.

Iron in cytochromes is alternatively oxidized (Fe+3) and reduced (Fe+2) .

The electrons are transported from coenzyme Q to cytochromes in the order b,c1,c, a and a3 during electron transport.

Components of ETCComponents of ETC

Complex-V ATP synthaseComplex-V ATP synthase

Enzyme that generates ATP sometimes referred as complex-V.

It has two functional subunits Fo (oha) and F1 portion

Chemi-osmotic theoryChemi-osmotic theoryThe proton pumps (complexes -I, III,

IV) expels H+ from inside to outside of the membrane.

So there is high H+ concentration outside .

This causes H+ to enter into mitochondria through the channels (Fo –F1complex ) , this proton influx binds to oxygen of pi+ADP to form ATP

Chemi-osmotic theoryChemi-osmotic theory

Around 3 protons are required per ATP synthesized.

When 1 NADH transfers its electrons to oxygen, 10 protons are pumped out this accounts for approximately 3 ATP synthesis.

Oxidation of 1 FADH2 is accompanied by the pumping of 6 protons accounting for 2 molecules of ATP.

Sites of ATP synthesisSites of ATP synthesis

Oxidative PhosphorylationOxidative Phosphorylation

The process of synthesizing ATP from ADP and Pi coupled with the electron transport chain is known as oxidative phosphorylation.

The complex V of the IMM is the site of O.P

Inhibitors of oxidative Inhibitors of oxidative phosphorylationphosphorylation1. Oligomycin: an antibiotic, used

as anti-fungal drug, prevents the cell from using the established H+-gradient, to make ATP.

2. Atractyloside: Plant toxin

Inhibitors of ETCInhibitors of ETCThe inhibitors bind to one of the

components of ETC and block the transport of electrons.

This causes the accumulation of reduced components before the blockade step and oxidized components after that step.

The synthesis of ATP is dependent on ETC.

Hence, all the site specific inhibitors of ETC also inhibit ATP formation.

Inhibitors of ETCInhibitors of ETC

Site- of inhibitorsSite- of inhibitors Site –I Between

complex-I and Co-Q:

1. Rotenone : a fish poison

2. Amytal : a barbiturate drug

3. Piercidin A: antibiotic

Site-II cytochrome Cytochrome reductase (b and c1):

1. Antimycin A : antibiotic

2. BAL (British antilewisite): an antidote used against war-gas

Site-III At complex-IV ( cytochrome oxidase):1. Carbon monoxide2. Cyanide3. Hydrogen Sulfide

UncouplersUncouplers

Compounds that can uncouple or delink the

electron transport chain from oxidative phosphorylation, such compounds are

known as Uncouplers.

The result is that ATP synthesis does not occur.

The energy linked with the transport of electrons is dissipated as heat.

Chemical UncouplersChemical Uncouplers1. 2,4-dinitrophenol

( has been extensively studied).

2. Dinitrocresol.3. Pentachlorophenol.4. Tri fluoro carbonyl

cyanide phenyl hydra zone (FCCP).

5. Aspirin (high doses).

Physiological UncouplersPhysiological Uncouplers

1. Thyroxine.2. Long chain fatty acids.3. Unconjugated Bilirubin. These act as Uncouplers only at

high concentration.

Significance of UncouplingSignificance of Uncoupling Brown adipose tissue

present in the upper back and neck portions and around kidney is rich in mitochondria and carry oxidation uncoupled from phosphorylation.

This causes liberation of heat when fat is oxidized in this tissue.

Significance of UncouplingSignificance of Uncoupling Examples :

1. New born infant, (Non-shivering Thermogenesis)

2. Hibernating animals

Brown adipose tissue is located in the neck area and is more physiologically active in the woman than in the man.

1. In certain individuals due to presence of this brown adipose tissue it is believed to protect them from becoming obese.

2. The excess calories consumed by this people are burnt and liberated as heat , instead of being stored as fat.

However, research, that was published in the New England Journal of Medicine, is showing that brown adipose tissue helps adults burn more calories than white adipose tissue.

Diseases associated with Diseases associated with oxida . phosoxida . phosLHON : Lebers hereditary optic

neuropathyMELAS: Mitochondrial

encephalopathy Lactic Acidosis Stroke

Learning check Learning check Mr. X 26yrs old male noted heat

intolerance, with profuse sweating . He is loosing weight in spite of good appetite . On physical Examination thyroid swelling was present and T3 and T4 are increased. Give biochemical explanation for above symptoms.