Chapter 7 Biological Oxidation. Biological oxidation is the cellular process in which the organic...

Chapter 7Chapter 7

Biological Biological OxidationOxidation

Biological oxidation is the cellular process in which the organic substances release energy (ATP), produce CO2 and H2O through oxidative-reductive reactions.

organic substances: carbohydrate, fat and protein

7.1 Principal of Redox Reaction7.1 Principal of Redox Reaction

The electron-donating molecule in a oxidation-reduction reaction is called the reducing agent or reductant;

the electron-accepting molecule is the oxidizing agent or oxidant:for example:

Fe2+ (ferrous) lose -e

Fe3+ (ferric) gain +e

Several forms of Biological OxidationSeveral forms of Biological Oxidation

1. Loss of electrons2. Dehydrogenation 3. Oxygenation

Redox reaction = reduction-oxidation reactionSeveral forms of Biological ReductionSeveral forms of Biological Reduction

1. Gain of electrons2. Hydrogenation 3. Deoxygenation

oxidation-reduction potential ( or redox potential), E : it is a measure of the affinity of a substance for electrons. It decide the loss (or the gain) of electrons.A positive E: the substance has a higher affinity for electrons , accept electrons easily.A negative E: the substance has a lower affinity for electrons , donate electrons easily.

E0`, the standard redox potential for a substance :is measured under stander condition(25℃, 1mmol/L reaction substance),at pH7, and is expressed in volts.

Section 7.2Section 7.2

Respiration Chain and Respiration Chain and Oxidative PhosphorylationOxidative Phosphorylation

7.2.1 Respiratory Chain 7.2.1 Respiratory Chain

•Term: A chain in the mitochondria consists o

f a number of redox carriers for transferring electrons from the substrate to molecular oxygen to form oxygen ion, which combines with protons to form water.

1.Complex I: NADH:ubiquinone oxidoreductase

NADH:CoQ oxidoreductase

2.Complex II: Succinate dehydrogenase

3.Complex III: cytochrome bc1 (ubiquinone Cyt c

oxidoreductase)4.Complex IV: cytochrome oxidase

Redox carriers including 4 protein complexes

Complex IComplex I ( (NADH:ubiquinoneNADH:ubiquinone oxidoreductaseoxidoreductase))

• Function: transfer electrons from NADH to CoQ

• Components: NADH dehydrogenase (FMN) Iron-sulfur proteins (Fe-S)

complex Ⅰ

NADH→ →Co

Q

FMN; Fe-SN-1a,b; Fe-SN-4; Fe-SN-3; Fe-SN-2

R=H: NAD+; R=H2PO3:NADP+

1. NAD(P)+: Nicotinamide Adenine Dinucleotide Phosphate)

Oxidation of NADH is a 2-Oxidation of NADH is a 2-electron(2e), 2-proton(2H) electron(2e), 2-proton(2H)

reactionreaction

NAD+ or NADP+ NADH or NADPH

2. FMN can transfer 1 or 2 hydride 2. FMN can transfer 1 or 2 hydride ions each timeions each time

Accepts 1 H+ and 1 e-

to form semiquinone = stable free radical

Accepts 2 H+ and 2 e-

to give fully reduced form

FMN: flavin mononucleotide

3. Iron-sulfur clusters (Fe-S) 3. Iron-sulfur clusters (Fe-S) transfers 1-electron at a time, transfers 1-electron at a time,

without proton involvedwithout proton involved

FeFe3+3++e+e- - Fe Fe2+2+

4.4.UbiquinoneUbiquinone ( (CoQCoQ) ) isis lipid-soluble lipid-soluble, not a c, not a component of complex omponent of complex ⅠⅠ,, can transfer 1 or 2 hydridcan transfer 1 or 2 hydrid

e ions each time.e ions each time. Function:Function: transfer electrons and protons from ctransfer electrons and protons from c

omplex Ⅰ,Ⅱto complex Ⅲ.omplex Ⅰ,Ⅱto complex Ⅲ.

NADH+H+

NAD+

FMN

FMNH2

Reduced Fe-S

Oxidized Fe-S

Q

QH2

MatrixMatrix

Intermembrane space

Complex IIComplex II:: SuccinateSuccinate dehydrogenasedehydrogenase ( (SuccinateSuccinate: CoQ oxid: CoQ oxid

oreductaseoreductase))• Function: transfer electrons from succina

te to CoQ• Components: Succinate dehydrogenase (FAD, Fe-S) Cytochrome b560

Complex ⅡSuccinate→ →CoQ Fe-S1; b560; FAD; Fe-S2 ; Fe-S3

Fe3++e- Fe3++e- Fe2+ Fe2+

Cytochromes a, b, c are heme proteins, their hCytochromes a, b, c are heme proteins, their heme irons participate redox reactions of e- traeme irons participate redox reactions of e- transportnsport..

Succinate

MatrixMatrix

Intermembrane space

Complex III:Complex III: cytochrome bc1cytochrome bc1 (ubiquinone Cyt c(ubiquinone Cyt c

oxidoreductase)oxidoreductase)

• Function: transfer electrons from CoQ to cytochrome c

• Components: iron-sulfur protein cytochrome b(b562, b566) cytochrome c1

complex Ⅲ QH2→ →Cyt c

b562; b566; Fe-S; c1

MatrixMatrix

Intermembrane space

Cytochrome c is soluble, which will transfer electrons to complex Ⅳ

Complex IVComplex IV:: cytochrome cytochrome oxidaseoxidase

• Function: transfer electrons from Cyt c to molecule oxygen, the final electron acceptor.

• Components: cytochrome aa3

copper ion (Cu2+) Cu2+ + e- Cu+

Complex IV

Cyt c → → O2

CuA→a→a3→CuB

Coenzyme Qubiquinone/ol

Cytochrome c

Sequence of respiratory Sequence of respiratory chainchain

Principles:• e- tend to flow from a redox pair with a lower

E°to one with a higher E°• In the e--transport chain, e--carriers are

arranged in order of increasing redox potential, making possible the gradual release of energy stored in NADH, FADH2

氧化还原对 Eº' (V)

NAD+/NADH+H+ -0.32

FMN/ FMNH2 -0.30

FAD/ FADH2 -0.06

Cyt b Fe3+/Fe2+ 0.04（或0.10）Q10/Q10H2 0.07

Cyt c1 Fe3+/ Fe2+ 0.22

Cyt c Fe3+/Fe2+ 0.25

Cyt a Fe3+ / Fe2+ 0.29

Cyt a3 Fe3+ / Fe2+ 0.55

1/2 O2/ H2O 0.82

呼吸链中各种氧化还原对的标准氧化还原电位Redox potential redox pair E0

There are two respiratory chains• NADH respiratory chain NADH Complex Ⅰ CoQ Complex Ⅲ cytochrome c Complex Ⅳ O2

• Succinate (FADH2) respiratory chain Succinate ComplexⅡ CoQ ComplexⅢ c

ytochrome c ComplexⅣ O2

NADH respiration

chain

FADH2

respiration chain

• The oxidation of organic nutritions produces the energy-rich molecules, NADH and FADH2.

• The oxidation of NADH or FADH2 in mitochondrial is the electron transferring through respiration chain.

• The free energy produced in electron transferring supports the phosphorylation of ADP to form ATP.

• The oxidation of NADH or FADH2 and the formation of ATP are coupled process, called Oxidation Phosphorylation.

7.2.2 Oxidative Phosphorylation7.2.2 Oxidative Phosphorylation

The Chemiosmotic TheoryThe Chemiosmotic Theory

• The free energy of electron transport is conserved by pumping protons from the mitochondrial matrix to the intermembrane space so as to create an electrochemical H+ gradient across the inner mitochondrial membrane. The electrochemical potential of this gradient is harnessed to synthesize ATP.

Peter Mitchell

EElectrochemical Hlectrochemical H++ gradient gradient ((Proton-motive forceProton-motive force))

2 components involved1. Chemical potential energy

due to difference in [H+] in two regions separated

by a membrane2. Electrical potential energy

that results from the separation of charge when a proton moves across the membrane without a electron.

Complex I:4 H+ expelled per e--pair transferred to Q

Complex III:4 H+ expelled pere--pair transferredto Cyt c

Complex IV:2e- + 2 H+ from matrix convert ½ O2 to H2O; 2 further H+ expelled from matrix

Conformation 1(high affinity for H+)

Conformation 2(low affinity for H+).

Proton pumping:Proton pumping: Reduction-Reduction-dependentdependent conformational switch ofconformational switch of an an e--transport complexe--transport complex

Inner

Membrane

ATP SynthaseATP SynthaseIntermembrane space

MatrixMatrix

(ab2c9-12)

(α3β3γδε)

C ring

Each of 3 -subunitscontains an active site

F1: multisubunitcomplex that catalyzesATP synthesis

F0 = proton-conductingtransmembrane unit

β-subunit take up ADP and Pi to form ATP

ADP + Pi ATP

H+ flow

β-subunit has three conformations:T (tight), L (loose), O (open)

When protons flow back through F0 channel, γ-subunit is rotated by the rotation of c ring, then the conformations of β-subunits are changed, this lead to the synthesis and release of ATP. To form a ATP need 3 protons flow into matrix.

F0

F1

胞液侧

基质侧

ATP4-

ADP3- H2PO4- ATP4- H+

H+

H2PO4-

H2PO4-

ADP3-

ADP3-

H+

H+

Intermembrane space

MatrixMatrix

Translocation of ATP , ADP and Pi.

P/O ratiosP/O ratios• P/O ratio is the rate of phosphate

incorporated into ATP to atoms of O2 utilized. It measure the number of ATP molecules formed per two electrons transfer through the respiratory chain.

• NADH respiratory chain : 2.5,

• FADH2 respiratory chain: 1.5

• During two electrons transfer through NADH respiratory chain, ten protons are pumped out of the matrix.

• To synthesis and translocation an ATP, four protons are needed.

• So, two electrons transport can result in 2.5 ATP.

• To succinate respiratory chain , two electrons transport can result in 1.5 ATP.

Regulation of Oxidative PhosphorylatRegulation of Oxidative Phosphorylationion

• 1.PMF (proton motive force) regulate the electron transport.

higher PMF lower rate of transport• 2.ADP concentration resting condition: energy demanded is l

ow, ADP concentration is low, the speed of Oxidative Phosphorytion is low.

active condition: the speed is high.

Inhibitor of Oxidative PhosphoInhibitor of Oxidative Phosphorylationrylation

• 1.Inhibitor of electron transport

××

××

××

Retonone Amytal

Antimycin ASuccinate Cyanide, AzideCarbon Monoxide

ⅠⅠ

Ⅳ Ⅳ

Cyt c

FF00

FF11

• 2.Uncoupling agents uncoupling protein (in brown adipose tissue), 2,4-dinitrophenol, Pentachlorophenol

HH+ +

HH+ +

ⅡⅡ

ⅢⅢ

Intermenbran space

Matrix

uncoupling protein

Q

H

+ H+

2,4-dinitrophnolADP+Pi ATP

heat

Intermembrane space

MatrixMatrixOligomycinOligomycin

C ring

3.Oligomycin bonds at the connection of F0 and F1, inhibit the function of ATP synthase.

OligomycinOligomycin

Ⅱ

Ⅰ Ⅲ Ⅳ

Succinate

Ⅴ

××

××

××××××

Uncoupling agent

Retonone Amytal Antimycin A

ATP and other Energy-rich compountsATP and other Energy-rich compounts

2P

OH

OH

OH

O O

OH

HHOH OH

HH

N

N

N

N

OO CHO

NH2

= p p

OH OH

ATPADP

AMP

～ ～

ATP has two energy-rich phosphoric acid anhydride bonds, the hydrolysis of each bond release more energy than simple phosphate esters.

Some Energy-rich compounds

ΔGº’Structure Exemple

phosphoenolpyruvate

creatine phosphate

acetyl phosphate

Acetyl CoA

• The hydrolysis of energy-rich bond: ΔGº’ = -5 ～ -15kcal/mol• The compounds with energy-rich bond ar

e high-energy compounds. • The hydrolysis of low-energy bond: ΔGº’ = -1 ～ -3kcal/mol • The compounds with low energy bond are

low-energy compounds.

Transport of high-energy bond Transport of high-energy bond energiesenergies

• 1.Substrate level phosphorylation Glycerate 1,3-biphosphate + ADP Glycerate 3-phosphate +ATP ΔGº’ = -4.5kcal/mol Phosphoenolpyruvate +ADP Pyruvate + ATP ΔGº’ = -7.5kcal/mol

~P~P

~P~P

ADP

ATP

Substrate level

phosphorylation

Oxidative

Phosphorylation Energy

utilization

2.ATP is the center of energy producing and utilizing.

3.Other nucleoside triphosphates are i3.Other nucleoside triphosphates are involved in energy transportnvolved in energy transport..

• GTP: gluconeogenesis protein synthesis• UTP: glycogen• CTP: lipid synthesis

4.Transport of the terminal 4.Transport of the terminal phosphate bond of ATP to the phosphate bond of ATP to the

other nucleosideother nucleoside

• Function of nucleoside diphosphate kinase

ATP + UDP ADP + UTP ATP + CDP ADP + CTP ATP + GDP ADP + GTP

• Function of adenylate kinase ADP + ADP ATP + AMP

7.3 Energy from cytosolic NAD7.3 Energy from cytosolic NADHH

• A mitochondrial NADH produce 2.5 ATP• A cytosolic NADH must be transported in

to mitochondrial for oxidation by two methods.

Glycerol phosphate shuttle 1.5 ATP Malate aspartate shuttle 2.5 ATP

Glycerol phosphate shuttle

FADH2

NAD+

FAD

Intermembranspace

Electron chain

dihydroxyacetone phosphate

PiCH2O-

CH2OH

C=O

PiCH2O-

CH2OH

C=O

Glycerol

phosphate

PiCH2O-

CH2OH

CHOH

PiCH2O-

CH2OH

CHOH

NADH+H+

Glycerol

phosphate

dihydroxyacetone

phosphate

Inner menbran

Glycerol

phosphate

dehydrogenase

Glycerol

phosphate

dehydrogenase

NADH +H+

NAD+

NADH +H+

NAD+

Malate α-ketoglutarate

carrier

Glutamate-aspartate carrier

-OOC-CH2-C-COO-

O

oxaloacetate

cytosol inner mitochondrial

membran matrix

Electron chain

Aspartate

-OOC-CH

2-C-COO

-

H3N

+

H

-OOC-CH2-C-COO-

OH

HMalate

Malate

Aspartate

Glutamate Glutamate

α-ketoglutarateα-ketoglutarate

oxaloacetate

Malate aspartate shuttle

7.4 Other Biological Oxidations7.4 Other Biological Oxidations

• Monoxygenases dioxygenase --add 2 atoms of O2

oxygenase to organic compounds. monoxygenase (mixed-function oxidase, hy

droxylase) --adds 1 oxygen atom to organic compound

s as a hydroxyl group.RH + NADPH + H+ + O2 ROH + NADP+ + H2O

The chief compounds of monoxygenase:

Cyt b5, Cyt P450, Cyt P450 reductase(FAD,FMN)

Free Radical Scavenging Free Radical Scavenging EnzymesEnzymes

Free Radical: the groups with an unpaired electron. (such as O2

﹣ 、 H2O2 、• OH)1.Superoxide dismutases(SODs)

2O2﹣+ 2H+ H2O2 + O2

SOD

H2O + O2

peroxidase

• 2.Glutathione peroxidase

GlutathionGlutathione peroxidase peroxidas

ee

H2O2

(ROOH)

H2O(ROH+H2O)

2G –SH

G –S – S – G

NADP+

Glutathione reductase

NADPH+H+

• 3.Catalase (in peroxisomes)

2H2O2 2H2O + O2

catalase

summarysummary