MEMBRANE-BOUND

ELECTRON TRANSFER AND

ATP SYNTHESIS (taken from

Chapter 18 of Stryer)

FREE ENERGY – MOST USEFUL

THERMODYNAMIC CONCEPT IN BIOCHEMISTRY

Living things require an input of free energy for 3 major

purposes

1. Mechanical – Muscle contraction and other cellular

movement

2. Active transport of molecules and ions

3. Synthesis of macromolecules and other biomolecules

from simple precursors

First law of thermodynamics

“Energy can be neither created nor

destroyed”

But, it can be converted from one

form into another

Free energy for these processes

comes from the environment

Phototrophs - obtained by trapping light energy

Chemotrophs – energy by oxidation of foodstuffs

Free energy donor for most energy requiring processes is

Adenosine triphosphate (ATP)

ATP

Large amounts of free energy is liberated when ATP is

hydrolysed to ADP + Pi or AMP + PPi

ATP is continuously formed and consumed

Phototrophs harness the free energy in light to generate ATP -

Photosynthesis

Chemotrophs form ATP by oxidation of fuel molecules –

Oxidative phosphorylation

OXIDATIVE

PHOSPHORYLATION

Glucose is converted to pyruvate

And under aerobic conditions undergoes oxidative

decarboxylation to form AcCoA which is then oxidised to

CO2 by the citric acid cycle

Stages of Catabolism

Citric Acid Cycle

Glycolysis

Activated Carriers

These pathways along with fatty acid oxidation produce

energy rich molecules NADH and FADH2 as well as small

amounts of ATP

Chemotrophs derive energy from oxidation of fuel molecules

and in aerobic organisms the ultimate electron acceptor is O2

Electron is not transferred directly

Electron is transferred through special carriers, Pyridine

nucleotides

Electron acceptor Electron donor

NAD+ NADH

FAD FADH2

Activated Carriers

Respiratory electron transfer is the transfer of electrons

from the NADH and FADH2 (formed in glycolysis, fatty

acid oxidation and the citric acid cycle) to molecular

oxygen, releasing energy.

Oxidative phosphorylation is the synthesis of ATP from

ADP and Pi using this energy.

Both processes are located on the IMM

Electron transfer and

oxidative phosphorylation

Mitochondrion

Outer membrane•Permeable (12000da)

•Porin – 30-35kd pore forming protein

Inner membrane•Impermeable all ions and polar molecules

•Possess family of transporter molecules (for

ATP/ADP , Pi , pyruvate, citrate , etc.).

•Matrix side (N-negative), cytosolic side (P-

postive)

Mitochondrion

Mitochondria are the result of an

Endosymbiotic event

Organelles contain their own DNA which encode 13

respiratory chain proteins

Many proteins encoded by cell nuclear DNA

Cells depend on organelle for oxidative phosphorylation ,

mitochondrion depend on cell for their very existence

Suggested that all extant mitochondria are derived from

bacterial Rickettsia prowazekii

Oxidative phosphorylation is conceptually

simple and mechanistically complex.

Flow of electrons from NADH and FADH2 to

O2 occurs via protein complexes located in the

IMM

Leads to the pumping of protons from the

matrix to the cytosol across the IMM.

ATP is synthesised when protons flow back

into the matrix via a protein complex in the

IMM.

Oxidative phosphorylation

An example of energy coupling via an

electrochemical gradient across a membrane.

REDOX POTENTIAL AND

FREE ENERGY CHANGES

The energy stored in ATP is expressed as the phosphoryl

transfer potential which is given by Go for hydrolysis of

ATP (-7.3kcal/mol)

The electron transfer potential of NADH is represented as Eo

the redox potential ( or reduction potential or oxidation-

reduction potential) which is an electrochemical concept.

Redox potential is measured relative to the H+: H2 couple

which has a defined redox potential of 0V (Volts).

Redox couples

X- + H+ X + 1

2H2

X- X + e-

H+ + e- 1

2H2

A negative redox potential means that a substance has a lower

affinity for electrons than H2 .

A positive redox potential means a substance has a higher

affinity for electrons than H2.

NAD+/ NADH at -0.32V is a strong reducing agent and

poised to donate electrons

1/2 O2/ H2O at +0.82V is a strong oxidising reagent and

poised to accept electrons.

1/2 O2 + NADH + H+ H2O + NAD+

The difference (Eo = 1.14V) is equivalent to -52.6

kcal/mole.

Redox potential

Electrons can be transferrred between groups that are

not in contact

Distance between electron carrying

groups

THE RESPIRATORY ELECTRON TRANSFER

CHAIN CONSISTS OF THREE PROTON PUMPS

LINKED BY TWO MOBILE ELECTRON

CARRIERS

Electrons are

transferred from

NADH to O2 by a

chain of three large

transmembrane

respiratory chain

protein complexes

I

II

III

IV

These are

a) Complex I also known as

NADH-Ubiquinone (UQ) oxidoreductase

NADH-Q reductase

b) Complex III also known as

Ubiquinol (UQH2)-Cytochrome c oxidoreductase

Cytochrome reductase

c) Complex IV also known as

Cytochrome c- Oxygen oxidoreductase

Cytochrome oxidase

Q- coenzyme Q or ubiquinone

NADH-Q Oxidoreductase

x 2e-

NADH-Q reductase

NADH transfer of e- to flavin

mononucleotide to produce FMNH2

e- from FMNH2 transferred to iron sulfur

clusters

e- from iron sulfur (Fe-S) clusters shuttle to

coenzyme Q (ubiquinone)

Results in pumping of 4 H+ out of matrix

NADH + Q + 5H+matrix NAD+ +QH2 + 4H+

cytosol

Succinate Q reductase

FADH2 already part of complex, transfers

electrons to Fe-S centres and then to Q

This transfer does not result in transport of

protons

Q-cytochrome c Oxidoreductase

Q-cytochrome c Oxidoreductase

Transfers e- from QH2 (2 e- ) cytochrome c

(1 e- ) via heme

Mechanism known as Q cycle

QH2 + 2Cyt cox + 2H+matrix Q +2Cyt cred + 4H+

cytosol

Cytochrome c Oxidase

Cytochrome c Oxidase

Proton transport by cytochrome c

oxidase

Electrons are carried from Complex I to Complex III

by UQH2, the hydrophobic quinol (reduced quinone)

diffuses rapidly within the IMM.

Electrons are carried from Complex III to Complex IV

by cytochrome c, a small hydrophilic peripheral

membrane protein located on the cytosolic or P side of

the IMM.

Complex II (Succinate-UQ oxidoreductase) is

membrane bound and contains the FADH2 as a

prosthetic group . So electrons from FADH2 feed in to

UQH2.

These respiratory chain complexes contain redox

groups to carry the electrons being transferred through

them. These are flavins, iron-sulfur clusters, haems

and copper ions.

PROTON PUMPS AND THE

ATP SYNTHASE

The free energy change of the reactions catalysed by

Complexes I, III and IV is large enough for them to

pump protons from the matrix or N side of the IMM

to the cytosolic or P side of the IMM.

There is not enough energy released in Complex II,

so no proton pumping occurs in this complex.

OXIDATION AND

PHOSPHORYLATION ARE

COUPLED BY A PROTON-

MOTIVE FORCEThis is the chemiosmotic hypothesis put forward by

Peter Mitchell in 1961.

Transfer of electrons from NADH (or FADH2) to

oxygen leads to the pumping of protons to the

cytosolic side of the IMM.

The H+ concentration (pH) becomes higher (lower

pH) on the cytosolic side, and an electrical

potential (membrane potential) with the cytosolic

side of the IMM positive is generated

So a proton-motive force (p) is generated which consists

of both a pH and a .

Mitchell proposed that this proton-motive force drives the

synthesis of ATP by another transmembrane protein

complex, as the protons return back across the IMM

through this protein complex.

This protein complex is called the ATPase (because like any

enzyme it is reversible and was first discovered by it’s

ability to hydrolyse ATP)

It’s preferred name is the ATP synthase.

ATPase

It is now thought that the proton-motive force induces a

conformational change in the ATP synthase, which allows

the release of tightly bound ATP (the product) from the

enzyme, and thus catalyses ATP synthesis.

So this is an example of energy coupling via an activated

protein conformation.

C-ring

Rotates 360º

Requires 3 protons

per molecule of

ATP formed

10 subunits

THE

COMPLETE

OXIDATION OF

GLUCOSE

YIELDS ABOUT

30 ATP

Net Yield per glucose

Glycolysis 2 ATP

Citric Acid cycle 2 ATP (GTP)

Oxidative phosphorylation ~26 ATP

Most of the ATP is generated by oxidative phosphorylation

POWER TRANSMISSION BY PROTON

GRADIENTS: A CENTRAL MOTIF OF

BIOENERGETICS

Proton gradients

power a variety

of energy-

requiring

processes i.e.

IT IS EVIDENT THAT PROTON GRADIENTS ARE A

CENTRAL INTERCONVERTIBLE CURRENCY OF FREE

ENERGY IN BIOLOGICAL SYSTEMS.

THE RATE OF OXIDATIVE PHOSPHORYLATION IS

DETERMINED BY THE NEED FOR ATP

Under most physiologic conditions, electron transfer is tightly

coupled to phosphorylation. Electrons do not usually flow

through the electron transfer chain unless ADP is

simultaneously phosphorylated to ATP.

Oxidative phosphorylation and thus electron transfer require a

supply of

NADH

O2

ADP and Pi

Regulation

The most important factor controlling the rate of oxidative

phosphorylation is the level of ADP

Regulated by the energy charge.

This regulation of the rate of oxidative phosphorylation by the

ADP level is called respiratory control.